METHODS FOR MODULATING TRANSCRIPTION FROM THE AMYLOID beta -PROTEIN PRECURSOR (APP) PROMOTER

Abstract: The application concerns methods for modulating transcription from the amyloid beta -protein precursor (APP) promoter. The upstreaam stimulatory factor (USF) is described as being capable of activating transcription from the APP promoter. Also described are USF binding compounds which are capable of down-regulating expression from the APP promoter. Preferred USF binding compounds are the amyloid precursor-like proteins APLP1 and APLP2. The application further concerns a screening assay for determining which candidate USF binding compounds are capable of causing down-regulation of transcription from the APP promoter.

Description: Methods for Modulating Transcription from the Amyloid β-Protein Precursor (APP) Promoter Part of the work performed during the development of this invention utilized U.S. Government Funds under NIH grants AG11899, NS/AG30428 and CA42567. The government may have certain rights to this invention. Field of the Invention The invention relates to methods for modulating transcription from the amyloid β-protein precursor (APP) promoter. Background of the Invention The amyloid β-protein precursor (APP) is a member of a highly conserved family of integral membrane glycoproteins, which currently includes APP and two APP-like proteins (APLP), APLPl and APLP2 (Wasco, W. et al. , Genomics 15 231-239 (1993); Wasco, W. et al , Nat. Genet. 5:95-100 (1993)). APP-like proteins have also been identified in mouse (Wasco, W. et al , Proc. Natl. Acad. Sci. USA 59:10758-10762 (1992); Slu , H. et al. , J. Biol. Chem. 269:2637-2644 (1994)), Drosophila (Rosen, D. et al. , Proc. Natl. Acad. Sci. USA 56:2478-2482 (1989)), and C. elegans (Daigle, I. et al. , Proc. Natl. Acad. Sci. USA 90:12045-12049 (1993)). However, only APP gives rise to the 4 kDa Aβ peptide that aggregates in senile plaques and cerebral blood vessel deposits in the brains of patients with Alzheimer's disease (Masters, C. L. et al. , EMBO J. 4:2751-2763 (1985); Glenner, G. G. et al. , Biochem. Biophys. Res. Commun. 720:885-890 (1984)). The accumulation of the Aβ peptide in amyloid plaques occurs in the brains of normal elderly individuals, to some extent, but is greatly enhanced in patients with Alzheimer's disease and Down's syndrome (Masters, C. L. et al, Proc. Natl. Acad. Sci. USA 82:4245- 4249 (1985)). The presence of a third copy of the APP gene on chromosome 21 in Down's syndrome patients and the subsequent increased levels of APP mRNA (Tanzi, R. E. et al. , Science 255:880-884 (1987); Rumble, B. et al. , New Engl. J. Med. 320:1446-1452 (1989)) suggest that overexpression of the APP gene most likely leads to amyloid deposition in these individuals. Altered regulation of APP transcription could conceivably lead to a similar situation in localized areas of brains of Alzheimer's disease patients. Experiments with transgenic mice showed that APP overexpression indeed leads to amyloid deposition (Quon et al, Nature 352:239 (1991); Wirac et al., Science 252:323 (1991)). Studies of the promoter region of the APP gene indicate that it lacks typical TATA and CAAT boxes and has multiple transcriptional start sites, characteristic of housekeeping genes (Salbaum, J. M. et al., EMBO J. 7:2807- 2813 (1988)). It has been reported that one or more elements located between position -94 and -35 are responsible for an 8-fold increase in gene expression in HeLa cells (Pollwein, P. et al. , Nucleic. Acids. Res. 20:63-68 (1992)). Studies of the mouse APP promoter have shown that two positive regulatory elements are located between positions -100 and -37, and that one of these elements binds the mouse Spl factor (Izumi, R. et al. , Gene 112: 189-195 (1992)). A "combined" element, in which the AP-4 site is followed by an overlapping AP-1 site (AP-l/AP-4 site) is situated approximately at position -45 and is completely conserved in the human, rat and mouse promoters (Izumi, R. et al , Gene ii2:189-195 (1992); Chernak, J. M., Gene 133:255-260 (1993)). The deletion of the region containing the AP-l/AP-4 site in the rat APP promoter causes a 30 % decrease in transcriptional activity in PC- 12 cells (Hoffman, P. W. et al. , Biochem. Biophys. Res. Commun. 201:610-617 (1994)). Interestingly, it has been shown that APP mRNA levels change dramatically during differentiation of embryonic P19 cells induced with retinoic acid (Fukuchi, K. et al., J. Neurochem. 55:1863-1873 (1992)). The presence of an unidentified factor interacting with the AP-l/AP-4 site of the APP gene has been reported in a variety of systems. In HeLa cells, it has been demonstrated that the Spl factor interacts with an upstream GC-rich region and competes for binding with a factor interacting with the AP-l/AP-4 site (Pollwein, P. et al. , Nucleic. Acids. Res. 20:63-68 (1992); Pollwein, P., Biochem. Biophys. Res. Commun. 190:637-647 (1993)). The binding of an unknown factor to the region containing the conserved AP-l/AP-4 site in the rat APP promoter has been reported to occur in PC-11 cells and in rat brain (Hoffman, P. W. et al, Biochem. Biophys. Res. Commun. 207:610-617 (1994)). Finally, Quitshke et al. used a DNA fragment lacking the upstream GC-rich region and found that an unidentified factor binds to the AP-l/AP-4 site of the APP promoter in Y79 cells (Quitschke, W. W. et al. , J. Biol. Chem. 267: 17362-17368 (1992)). This factor did not appear to be related to the known AP-1 or AP-4 transcriptional factors. However, the identity of the factor(s) which binds to the AP-l/AP-4 site of the APP promoter has not yet appeared in the literature. The identification of factors involved in transcriptional regulation of the APP gene would provide critical clues regarding the events leading to the formation of amyloid deposits. Summary of the Invention The invention provides methods for modulating transcription from the amyloid β-protein precursor (APP) promoter. The upstream stimulatory factor (USF) has been identified as the nuclear factor that specifically binds the AP- l/AP-4 site within the APP promoter. The inventors have discovered that, not only does USF bind to the APP promoter, but it also activate transcription. Thus, the invention provides methods for activating transcription from the APP promoter. The method involves activating transcription by binding the APP promoter with either native or recombinantly produced USF. The invention also provides methods for detecting transcription from the APP promoter. Detection methods can involve either expression of a "fusion" reporter protein or a primer extension analysis of mRNA transcripts. By the invention, the luciferase reporter protein is preferably used to detect transcription. The invention further provides methods for down-regulating transcription from the APP promoter. The methods involve contacting the USF transcriptional activator with a USF binding compound capable of interfering with USF binding to the AP-l/AP-4 site in the APP promoter. By the invention, depending on the relative amounts of USF and the USF binding compound that are present, transcription from the APP promoter can be modulated as desired. Candidate USF binding compounds which may down- regulate transcription from the APP promoter include polyclonal and monoclonal anti-USF antibodies, nucleic acid fragments which contain an E- Box element, and members of the helix-loop-helix (HLH) transcription factor family. Preferred USF binding compounds are APP itself and the amyloid precursor like proteins, APLPl and APLP2. Each of these compounds are capable of down-regulating transcription from the APP promoter. The invention further provides a screening assay for determining which USF binding compounds are capable of down-regulating transcription from the APP promoter. The method involves transfecting a host cell with a recombinant construct containing the APP promoter operably linked to a gene encoding a reporter protein; transfecting the host cell with a recombinant construct capable of expressing the USF protein; measuring reporter protein expression activated by USF binding to the APP promoter; transfecting the host cell with a recombinant construct either containing or capable of expressing a candidate USF binding compound; and measuring if a decrease in reporter protein expression is caused by the USF binding compound interfering with USF binding to the APP promoter. Brief Description of the Figures FIGURE 1A-B. DNA-protein interaction at the AP-l/AP-4 site. (A) Schematic representation of the APP proximal promoter region. The approximate position of the DK-1 fragment and the position of the overlapping AP-l/AP-4 sites within the DK-1 fragment are indicated. (B) EMSA performed with the DK-1 fragment using H4 nuclear extract. Competition was carried out with increasing amounts (0-50-fold) of unlabeled DNA fragments. Cold: DK-1 fragment. FIGURE 2. Competition assay using AP-1 and AP-4 consensus elements and a c-fos mAb. EMSA carried out with increasing amounts (0-50- fold) of competitors. Cold: DK-1 fragment. FIGURE 3. Competition assay using the USF core sequence EMSA, employing either labeled DK-1 fragment, with increasing amounts (0- 50-fold) of cold USF fragment, or labeled USF fragment with increasing amounts (0-50-fold) of cold DK-1 fragment. FIGURE 4A-B. Antiserum anti-USF binds the protein complex on the AP-l/AP-4 site. (A) Detection of USF in nuclear extracts obtained from H4 cells by Western blot analysis, using an antiserum raised against the 43 kDa form of USF. H4: 40 μg of nuclear extract from H4 cells. rUSF: 100 ng of recombinant 43 kDa USF. (B) Competition assay using a USF antiserum: EMSA in presence of increasing amounts of anti-USF antiserum, employing either labeled DK-1 fragment, or labeled AP-1 fragment. FIGURE 5. Interaction between recombinant USF and the AP-I/AP-4 site. EMSA using 70-700 pg of rUSF and labeled DK-1 fragment. EMSA employing rUSF, in presence of increasing amounts (0-50-fold) of either cold DK-1, or AP-l/AP-4 randomized fragments. FIGURE 6A-C. EMSA and cell-free transcription from the APP promoter with HeLa extracts. (A) EMSA showing the shift pattern with HeLa cell nuclear extracts, lμ H4 nuclear extract, rUSF: 700 pg rUSF. HeLa: 1 μg HeLa cell nuclear extract. (B) Determination of the 5 '-end of the in vitro transcription product of the hAPP-luciferase construct by primer extension and comparison with the sequencing product of the construct, using the same oligonucleotide. The major transcription initiation site is indicated by an arrow. (C) In vitro transcription in the presence of competing DNA fragments. 1 μg of the hAPP-luciferase and 0.5 g of the pMLΔ53(CA 2 T) templates were incubated in the absence and presence of a 20-fold molar excess of USF fragment or AP-l/AP-4 Random fragment. FIGURE 7. Transactivation of the APP promoter by USF in cell-free transcription assays. In vitro transcription assay with 1.5 μg of hAPP-luciferase and 1 μg pMLΔ53(CA 2 T) templates in the absence and presence of 10 and 100 ng rUSF. FIGURE 8. H4 neuroglioma cells were cotransfected with three constructs: the APP promoter-luciferase reporter gene construct, the USF expression vector or its control plasmid, and the APP/APLP family expression vectors or their control plasmid. The effects on expression from the APP promoter are shown graphically. FIGURE 9. Schematic representation of the mouse APLPl open reading frame and the relation of various cDNA clones. The 2361 base pair open reading frame and the non-coding region of the APLPl cDNA are shown. Also shown are the relative locations of two representative cDNA clones found in 11 libraries, 69A and 1A, and a clone obtained through the RACE procedure, J. Restriction enzyme sites: E = EcoRI, P = Pstl. FIGURE 10. Nucleotide and amino acid sequence of the APLPl cDNA. The composite nucleotide sequence (SΕQ ID NO: 15) and the predicted amino acid sequence (SΕQ ID NO: 16) of APLPl is shown. The predicted membrane spanning region is underlined. The location of the primers that were used for the RACE procedures are indicated by arrowed lines over the nucleotide sequences. The location of the peptide sequence used for the production of antisera is double underlined. Predicted N-glycosylation sites are underlined with a squiggly line and a region surrounding a potential tyrosine phosphorylation site is underlined by dots. The polyadenylation signal is indicated by bold face type and the stop codon is shown by an asterisk. FIGURE 11. Comparison of the APLPl (SEQ ID NO: 17) and APP (SEQ ID NO: 18) amino acid sequences. The UWGCG Bestfit analysis of the mouse APLPl and human APP 695 (Chen, et al, J. Biol. Chem. 265:3116- 3123 (1990)) is shown. Identities are indicated by a vertical line between the two amino acids. Similarities are indicated by a single or double dot. Gaps produced by the Bestfit alignment are shown by dots in the sequence. The βA4-protein sequence is underlined in the APP sequence. The identities are concentrated in three regions: APLP amino acids 21-211, 316-488, and 609- 654. FIGURE 12. Domains of homology. Regions of the amino acid sequences of the mouse amyloid precursor like protein (APLPl) (SEQ ID NOS: 19,22,25), the human amyloid precursor protein APP (SEQ ID NOS:20,23,26), the Drosophila amyloid precursor-like protein (APPL1) (SEQ ID NOS:21, 24,27) and the rat testis cDNA (testis) (SEQ ID NO:28) are compared. Amino acids that are identical in all of the sequences in the domain are shown as capital letters in bold face type and are identified by the presence of a vertical line (|) above the sequences. Amino acids that are the same in more than one sequence are shown as capital letters and have a dot (°) over the sequences. Amino acids that are not identical to any others are shown as lower case letters. The conserved cysteines are identified by the presence of a carrot underneath the sequence. Spans of particularly conserved amino acids are underlined. An N-glycosylation signal is identified by a double underline. Stop codons are indicated by an asterisk and the amino acid numbers of the sequences are shown at the beginning of each line. /15265 -8- FIGURE 13. Northern blots of mouse brain and neuroblastoma RNA. Poly A+ RNA (10 μg) from neuroblastoma (lane 1) and mouse brain (lane 2) was separated on agarose gels containing formaldehyde and transferred to nylon as described in Materials and Methods. The blot was probed with DNA corresponding to nucleotides 1482-1995 of the nucleotide sequence shown in FIGURE 10 (SEQ ID NO: 15). Sizes of hybridizing messages in kb are indicated. FIGURE 14A-C. Western Blots using antiserum 301 (See the Example). Mouse brain and neuroblastoma proteins were separated by 7.5% PAGE as described in Materials and Methods. (A) Mouse brain protein was probed with antiserum 301 or preimmune serum at a dilution of 1 : 100. The binding of antiserum 301 to the 65 and 33 kDa proteins is inhibited by the presence of increasing amounts of the peptide QQLRELQRH (SEQ ID NO:l), used to immunize the rabbit. Preimmune serum with no peptide (lane 1); immune serum with no peptide (lane 2); immune serum preabsorbed with 5 ng/ml peptide (lane 3); immune serum preabsorbed with 50 ng/ml of peptide (lane 4); immune serum preabsorbed with 500 ng/ml of peptide (lane 5). Preabsorption with 500 ng of an irrelevant yeast β-tubulin peptide had no effect on the binding (lane 6). (B) Neuroblastoma cell extracts probed with preimmune serum (lane 1) and 301 antiserum (lane 2). Both sera were used at a dilution of 1 :100. (C) Anti-peptide (QQLRELQRH) (SEQ ID NO:l) antiserum recognizes a β-galactosidase-APLPl fusion protein. Western blots on bacterially produced proteins. Lanes 1-3 were stained with preimmune serum from rabbit 301. Lanes 4-6 were stained with immune serum. Lanes 1 and 4: induced (temperature sensitive induction and promoter) cells containing a plasmid with its β-galactosidase gene fused to an APLPl cDNA fragment inappropriately oriented for production of an aΛPLPl epitope. Lanes 2 and 5: uninduced cells containing a plasmid with its β-galactosidase gene fused in frame to the APLP open reading frame. Lanes 3 and 6: same cells as in lanes 2 and 5 except induced. Induced cells were grown at 42°C. Uninduced cells were grown at 30°C. The arrowhead indicates a β-galactosidase-APLPl fusion protein recognized by immune serum but not by preimmune serum. That protein is approximately 24 kDa larger than β-galactosidase alone, as predicted, due to the insertion of 222 additional residues of APLPl open reading frame. FIGURE 15A-E. Immunofluorescence staining of mouse neuroblastoma cells with antiserum 301. Cells were stained with antiserum 301 at a dilution of 1:10,000 as described in Materials and Methods. Panel (A) shows neuroblastoma cells stained with antiserum 301. Panel (B) shows a higher magnification of a cell stained with antiserum 301 where the reticular pattern is evident. This staining pattern is similar to that seen when an antibody to a known Golgi enzyme (mannosidase II) is used to stain the cells (C). The perinuclear staining is competed by the addition of the peptide that was used as the antigen (D), and is not seen in the presence of preimmune serum (E). The magnification in a,c,d and e is 720X and in b 950X. FIGURE 16. Mapping of the APLPl locus using a somatic cell hybrid panel. All hybrids have been previously described (Brook et al, Hum. Genet. 87:65-72 (1991); Chartier-Harlin et al, Nature 553:884-846 (1991); and Geissler et al, Cell Mol. Genet. 77:197-214 (1991)). The portions of chromosome 19 retained in each human-rodent cell hybrid are illustrated, and the names of the representative cell lines are shown above. The presence (+) or absence (-) of APLPl in each hybrid cell line is indicated. FIGURE 17. Comparison of APLP2 (SEQ ID NO:29) and APP amino (SEQ ID NO: 30) acid sequences. An alignment of the human APLP2 amino acid sequence and the human APP695 (Kang et al , Nature 325:733-736 (1987)) was generated using the UWGCG GAP analysis. Gaps produced by the alignment are indicated by dots in the sequence. The location of the four PCR primers that were used to generate the SG190 probe are indicated by arrows above the amino acid sequence. Twelve conserved cysteines are indicated by carets under the sequence and a zinc-binding motif is indicated by a double underline. A conserved acidic-rich region is located between APLP2 amino acids 216 and 278. An N-glycosylation signal is underlined, an alternatively spliced exon is overlined, predicted transmembrane regions are shown in italics, and a clathrin binding motif is indicated by bold face type. Potential phosphorylation sites are indicated by a # sign (protein kinase C), a sign (caseine kinase I and II) or a ° sign (tyrosine kinase) over the sequence. Stop codons are indicated by an asterisk. FIGURE 18. Alignment of the amino acid sequences of the members of the APP-gene family (SEQ ID NOS :31-33). The sequences of human APLP2, mouse APLPl (Wasco et al. , Proc. Natl. Acad. Sci. USA 59:10758-10762 (1992)) are presented as aligned by the UWGCG PILEUP program. Amino acids that are not identical or conservatively substituted are indicated by a dash. Gaps in the amino acid sequence that were created by the alignment are shown by dots. The predicted initiator methionine for each protein is shown and stop codons are indicated by an asterisk. FIGURE 19 A-B. Distribution of human APLP2 gene transcripts. RNA from fetal human tissues (A) or adult human brain (B) was isolated, fractionated, transferred to nylon membranes, and hybridized with radiolabeled probe as previously described (Tanzi et al , Science 235:880-884 (1987)). (A) Hybridization of a PCR fragment corresponding to APLP2 amino acids 327 to 490 (APLP2) or a 3' 1.1 kb EcoRI APP cDNA fragment (FB63) were hybridized to RNA (20 μg) from human 20-22 week aborted fetal tissue obtained midtrimester under protocols approved by the institutional review board at Brigham and Women's Hospital. (B) Hybridization of the APLP2 PRC fragment or FB63 to RNA (10 μg) from adult human brain subregions: A10, frontal cortex; A17, striate cortex; A18, extrastriate cortex, A20, 21 temporal association cortex; A4, motor cortex; thalamus-VPL, thalamus-ventral posterolateral nucleus; A40, posterior perisylvian cortex-supramarginal gyri; A44, anterior perisylvian cortex-opercular gyri. Shown beneath panel B is a control hybridization with a glyceraldehyde-3-phosphate dehydrogenase cDNA (G3PD). The two autoradiograms are from independent hydribizations to the same filter. FIGURE 20. Northern blot of APLP2 to total RNA from normal and Down syndrome brains, adult normal and ADA cerebellum and frontal cortex. A PCR generated fragment corresponding to APLP2 amino acid 327 to 490 and FB63 (-APP) were hybridized to total RNA (25 μg) from 19-week normal (N) and Down syndrome (DS) brains, adult normal (N Cb) cerebellum, and adult normal (N FCtx) and AD (AD FCtx) frontal cortex. The two autoradiograms are from independent hybridizations to the same filter. FIGURE 21. Nonisotopic in situ localization of APLP2- oligonucleotide. A) In situ hybridization using a 45-mer specific for APLP2 (corresponding to amino acids 74-88 in FIGURE 17 (SEQ ID NO:29)) reveals staining of CA1 pyramidal neurons. The probe was end-labeled with biotin-21- dUTP using 3' terminal transferase and visualized by avidin-biotin-peroxidase reaction (Tanzi et al , Mol. Brain Res. :iτ. press; Hyman et al , Mol. Brain Res. ύi. press; Wasco et al. , Alzheimer's disease and related disorders 7992:selected communications (in press)), b) a negative control 45-mer corresponding to the other strand of the same region of APLP2 shows no significant staining. Magnification = 16X.3. Detailed Description of the Invention The present inventors recognized that the AP-l/AP-4 site in the amyloid β-protein precursor (APP) promoter contains a palindromic sequence which includes an E-box element. E-box sequences are binding sites for the helix- loop-helix (HLH) transcription factor family including myc and Myo-D, and are involved in the regulation of development and cell differentiation (Murre, C. et al, Cell 56:777-783 (1989); Tapscott, S. J. et al , Science 242:405-411 (1988)). In an attempt to identify the factor that binds to the AP-l/AP-4 element, the present inventors investigated a candidate mammalian activator, upstream stimulatory factor (USF), which interacts with the adenovirus major late promoter and to elements in several cellular gene promoters, including those for the insulin gene (Read, M. L. et al. , Biochem. J. 295:233-237 (1993)), the type I plasminogen activator inhibitor gene (Riccio, A. et al , Mol. Cell Biol. 72:1846-1855 (1992)) and the P53 tumor suppressor gene (Reisman, D. et al. , Nucleic. Acids. Res. 27:345-350 (1993)). The present inventors have discovered that USF binds specifically to the AP-l/AP-4 site in the APP promoter. Moreover, the inventors have discovered that USF is necessary to maintain elevated levels of APP mRNA and that recombinant USF elevates transcription levels from the APP promoter. This has been shown in a cell-free transcription system wherein exogenous USF was added to a construct containing the APP promoter and by transfecting a host cell containing the APP promoter with a construct encoding and capable of expressing USF. Thus, the present invention is directed to modulating transcription from the APP promoter by binding the promoter with either exogenously added or recombinantly produced USF. The method involves binding the AP-l/AP-4 site in the APP promoter with either native or rUSF whereby transcription from the promoter is activated. Native and recombinant USF are readily available. Native USF can be obtained from H4 neuroglioma cells or HeLa cells by preparing nuclear extracts as described in Miner et al, J. Neurosci. Res. 33:10 (1992). Alternatively, cDNA encoding the 43 kDa form of USF has been cloned, sequenced and expressed in bacteria (Gregor et al. Genes. Dev. 4.1730 (1990)). It has been shown that the recombinantly expressed 43-kDa USF binds to its cognate DNA sequence in a manner indistinguishable from that of HeLa USF (Pognonec & Roeder, Molecular and Cellular Biology 11:5125 (1991)). It would be well within the ability of the skilled artisan to subclone the gene encoding USF into an appropriate DNA or RNA expression vector depending on the host cell to be transfected and the level of expression sought. For example, the present inventors have subcloned the USF gene into a retroviral vector expression system and transfected H4 neuroglioma cells with the resulting construct. The control sequences, expression vectors, and transformation methods are dependent on the type of host cell used to express the USF gene. Generally, procaryotic, yeast, or mammalian cells are useful as hosts. Procaryots most frequently are represented by various strains of E. coli. However, other microbial strains may also be used, such as bacilli, for example Bacilllus subtilis, various species of Pseudomonas, or other bacterial strains. In such procaryotic systems, plasmid or bacteriophage vectors which contain replication sites and control sequences derived from a species compatible with the host are used. A wide variety of vectors for many procaryotes are known (Maniatis et al, (1982) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Sambrook et al, Molecular Cloning (1989) Molecular Cloning: A Laboratory Manual, 2nd Ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Methods of Enzymology Vol. 68, 100, 101, 152-155, Academic Press, Orlando (1979, 1983, 1987). Commonly used procaryotic control sequences include promoters for transcription initiation, optionally with an operator, along with ribosome binding site sequences, include such commonly used promoters as the beta- lactamase (penicillinase) and lactose (lac) promoter systems, the tryptophan (trp) promoter system and the lambda derived PL promoter and N-gene ribosome binding site, which has been made useful as a portable control cassette (U.S. Pat. No. 4,711,845). However, any available promoter system compatible with procaryotes can be used. In addition to bacteria, eucaryotic microbes, such as yeast, may also be used as hosts for the expression of USF. Laboratory strains of Saccharomyces cerevisiae, Baker's yeast, are most used although a number of other strains are commonly available. Vectors employing the 2 micron origin of replication and, other plasmid vectors suitable for yeast expression are known (Maniatis et al, (7952; Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Sambrook et al, Molecular Cloning (1989) Molecular Cloning: A Laboratory Manual, 2nd Ed. , Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Methods of Enzymology , Vol. 68, 100, 101, 152-155, Academic Press, Orlando (1979, 1983, 1987); Pouwels et al , Cloning Vectors: A Laboratory Manual. Elsevier, Amsterdam (1987)). Control sequences for yeast vectors include promoters for the synthesis of glycolytic enzymes. Additional promoters known in the art include the promoter for 3- phosphoglycerate kinase and those for other glycolytic enzymes, such as glyceraldehyde-3-phosphate dehydrogenase, hexokinase, pyruvate decarboxylase, phosphofructokinase, glucose-6-phosphate isomerase, 3- phosphoglycerate mutase, pyruvate kinase, triosephosphate isomerase, phosphoglucose isomerase, and glucokinase. Other promoters, which have the additional advantage of transcription controlled by growth conditions, are the promoter regions for alcohol dehydrogenase 2, isocytochrome C, acid phosphatase, degradative enzymes associated with nitrogen metabolism, and enzymes responsible for maltose and galactose utilization. It is also believed that terminator sequences are desirable at the 3' end of the coding sequences. Such terminators are found in the 3' untranslated region following the coding sequences in yeast derived genes. Many of the vectors illustrated contain control sequences derived from the enolase gene containing plasmid peno-46 or the LEU2 gene obtained from YEpl3, however, any vector containing a yeast compatible promoter, origin of replication and other control sequences is suitable (Maniatis et al, (1982) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Sambrook et al, Molecular Cloning (1989) Molecular Cloning: A Laboratory Manual, 2nd Ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Methods of Enzymology , Vol. 68, 100, 101, 152-155, Academic Press, Orlando (1979, 1983, 1987); Pouwels et al , Cloning Vectors: A Laboratory Manual. Elsevier, Amsterdam (1987)). It is also possible to express the gene encoding USF in eucaryotic host cells from multicellular organisms (Freshly, R.I., Culture of Animal Cells: A Manual of Basic Technique, 2nd Ed., Alan R. Liss, New York (1987)). Useful host cell lines include murine myelomas N51, VERO, and HeT cells, and Chinese hamster ovary (CHO) cells. Expression vectors for such cells ordinarily include promoters and control sequences compatible with mammalian cells such as, for example, the commonly used early and late promoter from Simian Virus 40 (SV40), or other viral promoters such as those derived from polyoma, Adenovirus 2, bovine papilloma virus, or avian sarcoma viruses, or immunoglobulin promoters and heat shock promoters (Maniatis et al, (1982) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Sambrook et al, Molecular Cloning (1989) Molecular Cloning: A Laboratory Manual, 2nd Ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Methods of Enzymology , Vol. 68, 100, 101, 152-155, Academic Press, Orlando (1979, 1983, 1987); Pouwels et al, Cloning Vectors: A Laboratory Manual. Elsevier, Amsterdam (1987)). General aspects of mammalian cell host system transformations have been described by aAxel (U.S. Patent No. 4,399,216). Origins of replication may be obtained, if needed, from viral sources. However, integration into the chromosome is a common mechanism for DNA replication in eucaryotes. Plant cells are now also available as hosts, and control sequences compatible with plant cells such as the nopaline synthase promoter and polyadenylation signal sequences are available (Pouwels et al, Cloning Vectors: A Laboratory Manual. Elsevier, Amsterdam (1987); Methods of Enzymology, Vol 118, Academic Press, Orlando (1986); Gelvin et al, Plant Molecular Biology Manual, Kluwer Academic Publishers, Dudrecht (1990)). Depending on the host cell used, transformation is done using standard techniques. Such techniques include calcium treatment employing calcium chloride for procaryotes or other cells which contain substantial cell wall barriers; infecting with Agrobacterium tumefaciens for certain plant cells; calcium phosphate precipitation method for mammalian cells without cells walls; and, macroprojectile bombardment for many cells including plant cells. The present inventors have shown that transcription of a reporter protein from an APP promoter present in H4 neuroglioma cells can be enhanced 5-fold by transfecting the cells with a retroviral expression vector capable of expressing USF. Thus, it will be recognized that a variety of expression vectors can be used for expressing the USF gene depending on what type of cells contain the APP promoter. Delivery of the gene encoding USF to cells in humans for modulating APP expression can occur using one of the many known vector systems known in the art. Retroviral vectors can only integrate into the genome of dividing cells. Thus, these vectors provide a useful vehicle for the selective targeting of USF to dividing cells. Retroviral vectors offer further advantages as there are no limitations in host range and these vectors have already been used successfully to infect many different cell types (Cepko, C, "Lineage analysis and immortalization of neural cells vial retrovirus vectors", in Neuromethods, Vol. 16, 177-218, Clifton, NJ, The Humana Press, Inc. (1989); Gilboa, E., BioEssays 5(6):252-257 (1987); Friedmann, T., Science 244: 1275-1281 (1989)). In general retroviral expression vectors that are effective for integrating genes into dividing cells are well known in the art (Breakfield et al, Molec. Neuro. Biol. 1:229 (1987); Breakefiled et al. The New Biologist 3:203 (1991); Huang et al, Experimental Neurology 775:303 (1992), WO93/03743 and WO90/09441. APP is expressed in all major tissues (Schmechel et al, Alzheimer Dis. Assoc. Disord. 2:96 (1988)). In the brain, APP is expressed primarily, but not exclusively, in neurons (Schmechel et al, Alzheimer Dis. Assoc. Disord. 2:96 (1988)). Expression vectors capable of infecting neurons are known and include those based on the Herpes Simplex Virus. The APP promoter has been cloned and sequenced (Quitschke & Goldgaber, The Journal of Biological Chemistry 267:17362 (1992)). The inventors have discovered that USF will bind to DNA fragments containing the AP-l/AP-4 element located in the APP promoter. Example 1 of the specification shows that the sequence of the DK-1 fragment, which extends from -30 to -58 from the primary transcriptional site of the human APP promoter (Salbaum, J. M. et al. , EMBO J. 7:2807-2813 (1988)) and contains the AP-l/AP-4 element (underlined): 5'GGGCCGGATCAGCTGACTCGCCTGGCTCT'3 (SEQ ID NO:2), is specifically bound by both native and rUSF. Preferably, the APP promoter is operably linked to a nucleic acid sequence encoding either the APP protein or a heterologous polypeptide. Of course, the APP promoter is operably linked to the APP gene in the variety of cells which naturally express APP. In another preferred embodiment, the APP promoter is operably linked to a nucleic acid sequence encoding a reporter protein. Such "fusion" reporter proteins are well known in the art. A reporter gene encoding a reporter protein may be fused to the APP promoter or fragments thereof containing the AP-l/AP-4 element. The amount of reporter gene product produced is indicative of the relative activity of the promoter. Thus, the present invention is further directed methods for detecting transcription from the APP promoter. One example of a suitable reporter protein is β-galactosidase. β- galactosidase is an enzyme encoded by the lac Z gene of E. coli. The presence of the lac Z gene product in a cell can be qualitatively determined in whole cells and can be quantitatively measured in cell-free extracts. Other reporter genes that can be used include: β-galactosidase (MacGregor et al. , 1987, Somatic Cell Mol. Genet. 73:253-266); galactokinase (e.g., Rosenberg et al , 1983, Science 222:734-739; McKenney et al. , 1981 , in Gene Amplification and Analysis 2:383-415, Elsevier/North-Holland, New York); Murooka and Mitani, 1985, /. Biotechnol 2:303-316; β-glucuronidase (e.g., Jefferson et al. , 1986, Proc. Natl. Acad. Sci. USA 53:8447-8541); human growth hormone (e.g., Seldon et al. , 1986, Mol. Cell. Biol. 6:3173-3179); chloramphenicol acetyltransferase (CAT) (e.g., Tsukada et al., 1987, J. Biol. Chem. 262:8743- 8747; Carbonell and Miller, 1987, Appl. Environ. Microbiol. 53:1412-1417; Boulet et al. , 1986, Proc. Natl. Acad. Sci. USA 53:3599-3603; Jameson et al. , 1986, Endocrinology 779:2560-2567; Montminy et al. , 1986, Proc. Natl. Acad. Sci. USA 53:6682-6686); Tn5 neomycin phosphotransferase (e.g., Kaulen et al. , 1986, EMBO J. 5: 1-8; Simpson et al. , 1985, EMBO J. 4:2723- 2730) and firefly luciferase (e.g., Ow et al., 1987, Proc. Natl. Acad. Sci. USA 54:4870-4874, Ow et al , 1986, Science 234:856-859). Quitschke et al. fused the APP promoter 5' and operably linked to the reporter gene chloramphenicol acetyltransferase (Quitschke et al, The Journal of Biological Chemistry 267(24) : 17362 (1992)). The present inventors have fused the APP promoter 5' and operably linked to the luciferase reporter gene. The luciferase reporter gene system is discussed in detail in U.S. Patent No. 5,196,424. An alternative method for detecting transcription from the APP promoter involves primer extension analysis of the resulting transcript. Primer extension analysis involves hybridizing a primer having a sequence complementary to a portion of the transcribed mRNA and performing primer extension. Preferably, the primer is detectably labelled. Primer extension analysis is well known in the art. The present inventors, for example, prepared an APP promoter-luciferase reporter gene construct by inserting a 2.9 EcoRI/BamHI APP promoter fragment (Salbaum et al, EMBO J. 7:2807 (1988)) into the Smal site of the luciferase expression vector pxP2 (Nordeen, S.K., Biotechniques 6:454 (1988)). Cell-free in vitro transcription of the construct was performed according to the method of Dignam et al, Nucleic Acids Res 77:1475 (1983). Transcription from the construct was then detected by hybridizing a primer corresponding to part of the multiple cloning site between the APP promoter and the reporter gene. The primer had the following sequence: 5'-GCTCAGATCTCGAGCTCGGTAC-3' (SEQ ID NO: 3). There are a variety of compounds which bind USF. These include anti- USF polyclonal and monoclonal antibodies, nucleic acid fragments which contain E-Box sequences (CANNTG), and helix-loop-helix (HLH) transcription factors. The present inventors have discovered that certain USF binding compounds are capable of interfering with USF binding to the APP promoter thereby down-regulating transcription. By "down-regulating transcription from the APP promoter" is intended reducing transcription from the APP promoter relative to the transcription level attained in the absence of the USF binding compound. Thus, a further aspect of the present invention is directed to down- regulating transcription from the APP promoter using one or more USF binding compounds. The method involves contacting the USF transcription activator with a USF binding compound(s) capable of interfering with USF binding to the APP promoter. Depending on the relative amounts of USF and the USF binding compound mat are present, transcription from the APP promoter can be modulated as desired. For example, the present inventors transfected host cells with the APP promoter-luciferase reporter gene construct discussed above. The cells were then transfected with an expression vector encoding the USF gene (pCMV-USF). This activated transcription from the APP promoter about 5-fold relative to the transcription level attained in the presence of a control plasmid not encoding the USF transcription activator. The cells were then transfected with an expression vector encoding one of the following USF binding compounds: APLPl, APLP2, and APP. The results showed that the presence of each member of the APP/APLP family decreases the level of USF activation of transcription from the APP promoter at least 50%. As indicated, candidate USF binding compounds which may down- regulate transcription from the APP promoter include, but are not limited to, polyclonal and monoclonal anti-USF antibodies, nucleic acid fragments which contain E-Box sequences (CANNTG), and members of the helix-loop-helix (HLH) transcription factor family. Polyclonal and monoclonal antibodies can be raised against USF according to conventional techniques. For example, USF antiserum can be prepared as described in Kaulen et al, Mol. Cell. Biol. 77:412 (1991). Using an electrophoretic mobility shift assay (EMSA), the present inventors have shown that USF antiserum causes a marked decrease in the amount of USF complex formation with the AP-l/AP-4 site in the APP promoter. Nucleic acid fragments which contain an E-Box sequence can be any fragment containing the sequence CANNTG (or CANNUG if the fragment comprises RNA), where "N" is a nucleotide containing any of the four bases: Adenine, Guanine, thymidine (Uracil), or Cytosine. Preferably, "N" is a nucleotide containing either Cytosine or Guanine. For example, the core sequence of the AP-l/AP-4 site is CAGCTG. USF also binds the core sequence of the major late adenoviral promoter, CACGTG. By the invention, exogenously added or recombinantly produced nucleic acid fragments containing an E-Box sequence down-regulate transcription from the APP promoter through competition for USF binding. For example, in competition assays, the present inventors have shown that the presence of a DNA fragment containing an E-Box sequence significantly decreases transcription from the APP promoter. Conversely, the presence of the same DNA fragment in which the E-Box sequence was replaced by a random sequence does not decrease transcription. Whether any given DNA fragment containing an E-Box sequence is capable of down-regulating transcription from the APP promoter can easily be determined empirically. USF is a basic helix-loop-helix (HLH) protein, binding DNA through the basic region, and other HLH proteins through the HLH domain. The APP/APLP family of proteins contain a region homologous to an HLH domain, but the preceding region is only slightly basic. Other HLH proteins which display features similar to that of the APP/APLP family have been characterized and have been shown to bind to basic HLH proteins other than USF. For example, the protein called Id specifically represses transcriptional activation by the basic HLH myogenic factor, MyoD. It is these proteins which display features similar to that of the APP/APLP family (i.e., which modulate transcriptional activation by basic HLH proteins) that are candidate USF binding compoimds. Such candidate proteins, or peptide fragments thereof, can be screened for the ability to down-regulate transcription from the APP promoter using the screening assay described below. The present inventors have discovered that amyloid precursor-like proteins (APLP) and APP itself are capable of down-regulating transcription from the APP promoter by interfering with USF binding to the AP-l/AP-4 site. By "amyloid precursor-like proteins" (APLP) is intended to include any amyloid precursor-like protein, from any species, including APLPl and APLP2, especially from human brain, Alzheimer's disease human brain, or a synthetic APLP. The present APLP exhibits at least 40% identity at the amino acid level to APP and or APLPl and/or APLP2, more preferably at least 50% identity thereto, and contains an N-terminal cysteine-rich region consisting of at least 10 cysteines, more preferably consists of 12 cysteines. The term is also used in this invention to include any analog, homolog, mutant or derivative of a naturally occurring APLP. The term is also meant to include fragments having less than the naturally occurring number of amino acids, such as partial fragments of natural or synthetic APLP which retain the biological or immunological characteristics of the polypeptides specifically disclosed in this application. The term is also used to include any product which comprises the sequence of a naturally occurring APLP, or analog or homolog thereof, together with one or more flanking amino acids, which still have the same biological or immunological characteristics. The term is also used to include any peptide which comprises the sequence of a naturally-occurring APLP or an analog thereof together with one or more flanking amino acids, which have the same biological (functional or structural) or immunological characteristics. -22- The APLP's suitable for use in the present invention can be administered exogenously or by expression using recombinant DNA techniques. Methods for isolating, cloning, and recombinantly expressing APLP's are discussed below. As indicated, the present invention is further directed to a screening assay for identifying which USF binding compounds are capable of down- regulating transcription from the APP promoter. The method involves transfecting a host cell with a DNA or RNA construct containing the APP promoter operably linked to a gene encoding a reporter protein; transfecting the host cell with a DNA or RNA construct capable of expressing the USF protein; measuring reporter protein expression activated by USF binding to the APP promoter; transfecting the host cell with a DNA or RNA construct either containing or capable of expressing a USF binding compound; and measuring if a decrease in reporter protein expression is caused by the USF binding compound interfering with USF binding to the APP promoter. Preferably, the gene encodes luciferase reporter protein, the DNA binding compound is an APLP, and the host cells are neurogliomas. More preferably, the APLP is APLP-1 or APLP-2 or fragments thereof capable of decreasing transcription. Techniques for measuring luciferase reporter protein levels are well known in the art (See, for example, U.S. Patent No. 5,196,424). Techniques for measuring levels of other reporter proteins are also known in the art (See the reference cited above). Due to its simplicity, the screening method of the present invention is suitable for screening large numbers of USF binding compounds to determine which are capable of reducing transcription from the APP promoter. A detailed description of methods for isolating, cloning, and expressing APLP proteins is provided below and in co-pending application no. 08/007,999, which is herein incorporated by reference. Having established the amino acid sequence of both APLPl and APLP2, a nucleotide probe can be constructed which is complementary to the DNA, or mRNA coding for APLPl or APLP2 or a fragment thereof. This probe can be used as a diagnostic test to determine the presence of other APLPs. The process for genetically engineering APLPl and APLP2 sequences, is facilitated through the cloning of genetic sequences which are capable of encoding the peptide and through the expression of such genetic sequences. Genetic sequences which are capable of encoding the present APLP proteins are derived from a variety of sources. These sources include genomic DNA, cDNA, synthetic DNA, and combinations thereof. The preferred source of the genomic DNA or mRNA is brain or neuroblastoma cells. Post mortem RNA procedures can be followed to isolate the RNA. See Sajdel-Sulkowska et al, J. Neurochem. 40:670-680 (1983). The mRNA may then be used to obtain cDNA by techniques known to those skilled in the art. Probes may be synthesized based on the known amino acid sequence of the present APLP proteins (APLPl and APLP2) by methods known in the art. APLP mRNA can be isolated from any cell which produces or expresses APLP, and used to produce cDNA by means well known in the art (for example, see Guide to Molecular Cloning Techniques, S.L. Berger et al. , eds., Academic Press (1987)). Preferably, the mRNA preparation used will be enriched in mRNA coding for APLP, either naturally, by isolation from cells which are producing large amounts of die protein, or in vitro, by techniques commonly used to enrich mRNA preparations for specific sequences, such as sucrose gradient centrifugation, or both. For cloning into a vector, such suitable DNA preparations (either human genomic DNA or cDNA) are randomly sheared or enzymatically cleaved, respectively, and ligated into appropriate vectors to form a recombinant gene (either genomic or cDNA) library. A DNA sequence encoding APLP or its functional derivatives may be inserted into a DNA vector in accordance with conventional techniques, including blunt-ending or staggered-ending termini for ligation, restriction enzyme digestion to provide appropriate termini, filling in of cohesive ends as appropriate, alkaline phosphatase treatment to avoid undesirable joining, and ligation with appropriate ligases. Techniques for such manipulations are disclosed by Maniatis et al, (In: Molecular cloning, A Laboratory Manual, Cold Spring Harbor Laboratories, Cold Spring Harbor, NY, (1982)), and are well known in the art. Libraries containing APLP clones may be screened and an .APLP clone identified by any means which specifically selects for APLP DNA such as, for example, a) by hybridization with an appropriate nucleic acid probe(s) containing a sequence specific for the DNA of this protein, or b) by hybridization-selected translational analysis in which native mRNA which hybridizes to the clone in question is translated in vitro and the translation products are further characterized, or, c) if the cloned genetic sequences are themselves capable of expressing mRNA, by immunoprecipitation of a translated APLP product produced by the host containing the clone. Oligonucleotide probes specific for APLP which can be used to identify clones to this protein can be designed from knowledge of the amino acid sequence of APLPl or APLP2. Because the genetic code is degenerate, more than one codon may be used to encode a particular amino acid (Watson, J.D., In: Molecular Biology of the Gene, 3rd Ed., W.A. Benjamin, Inc., Menlo Park, CA (1977), pp. 356- 357). The peptide fragments are analyzed to identify sequences of amino acids which may be encoded by oligonucleotides having the lowest degree of degeneracy. This is preferably accomplished by identifying sequences that contain amino acids which are encoded by only a single codon. Although occasionally an amino acid sequence may be encoded by only a single oligonucleotide sequence, frequently the amino acid sequence may be encoded by any of a set of similar oligonucleotides. Importantly, whereas all of the members of this set contain oligonucleotide sequences which are capable of encoding the same peptide fragment and, thus, potentially contain the same oligonucleotide sequence as the gene which encodes the peptide fragment, only one member of the set contains the nucleotide sequence that is identical to the exon coding sequence of the gene. Because this member is present within the set, and is capable of hybridizing to DNA even in the presence of the other members of the set, it is possible to employ the unfractionated set of oligonucleotides in the same manner in which one would employ a single oligo- nucleotide to clone the gene that encodes the peptide. Using the genetic code (Watson, J.D., In: Molecular Biology of the Gene, 3rd Ed., W. A. Benjamin, Inc., Menlo Park, CA (1977)), one or more different oligonucleotides can be identified from the amino acid sequence, each of which would be capable of encoding the APLP. The probability that a particular oligonucleotide will, in fact, constitute the actual APLP coding sequence can be estimated by considering abnormal base pairing relationships and the frequency with which a particular codon is actually used (to encode a particular amino acid) in eukaryotic cells. Such "codon usage rules" are disclosed by Lathe et al. , J. Molec. Biol. 753:1-12 (1985). Using the "codon usage rules" of Lathe, a single oligonucleotide sequence, or a set of oligonucleotide sequences, that contain a theoretical "most probable" nucleotide sequence capable of encoding the APLP sequences is identified. The suitable oligonucleotide, or set of oligonucleotides, which is capable of encoding a fragment of an APLP gene (or which is complementary to such an oligonucleotide, or set of oligonucleotides) may be synthesized by means well known in the art (see, for example, Synthesis and Application of DNA and RNA, S.A. Narang, ed., 1987, Academic Press, San Diego, CA) and employed as a probe to identify and isolate the cloned APLP gene by techniques known in the art. Techniques of nucleic acid hybridization and clone identification are disclosed by Maniatis et al. (In: Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratories, Cold Spring Harbor, NY (1982)); Berger et al. , (In: Guide to Molecular Cloning Techniques, Academic Press (1988)); Sambrook et al. , (In: Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Press, Cold Spring Harbor, NY, 2d ed. (1989); and by Hames et al. (In: Nucleic Acid Hybridization, A Practical Approach, IRL Press, Washington, DC (1985)), which references are herein incorporated by reference. Those members of the above-described gene library which are found to be capable of such hybridization are then analyzed to determine the extent and nature of the APLP encoding sequences which they contain. To facilitate the detection of die desired APLP DNA encoding sequence, the above-described DNA probe is labeled with a detectable group. Such detectable group can be any material having a detectable physical or chemical property. Such materials have been well-developed in the field of nucleic acid hybridization and in general most any label useful in such methods can be applied to the present invention. Particularly useful are radioactive labels, such as 32 P, 3 H, I4 C, 35 S, l2S l, or the like. Any radioactive label may be employed which provides for an adequate signal and has a sufficient half-life. The oligonucleotide may be radioactively labeled, for example, by "nick- translation" by well-known means, as described in, for example, Rigby et al , J. Mol. Biol. 773:237 (1977) and by T4 DNA polymerase replacement synthesis as described in, for example, Deen et al. , Anal. Biochem. 735:456 (1983). Alternatively, polynucleotides are also useful as nucleic acid hybridization probes when labeled widi a non-radioactive marker such as biotin, an enzyme or a fluorescent group. See, for example, Leary et al. , Proc. Natl. Acad. Sci. USA 50:4045 (1983); Renz et al, Nucl. Acids Res. 12:3435 (1984); and Renz, M., EMBO J. 6:817 (1983). Thus, in summary, the actual identification of APLP sequences permits the identification of a Uieoretical "most probable" DNA sequence, or a set of such sequences, capable of encoding such a peptide. By constructing an oligonucleotide complementary to this theoretical sequence (or by constructing a set of oligonucleotides complementary to the set of "most probable" oligo¬ nucleotides), one obtains a DNA molecule (or set of DNA molecules), capable of functioning as a probe(s) for the identification and isolation of clones containing an APLP (i.e. APLPl or APLP2) gene. In an alternative way of cloning an APLP gene, a library is prepared using an expression vector, by cloning DNA or, more preferably cDNA prepared from a cell capable of expressing an APLP, into .an expression vector. The library is men screened for members which express the APLP, for example, by screening the library with antibodies to the protein. The above discussed methods are, therefore, capable of identifying genetic sequences which are capable of encoding an APLP or fragments of an APLP protein. In order to further characterize such genetic sequences, and, in order to produce the recombinant protein, it is desirable to express the proteins which these sequences encode. Such expression identifies diose clones which express proteins possessing characteristics of an APLP. Such characteristics may include the ability to specifically bind an APLP antibody and the ability to elicit the production of antibody which are capable of binding to an APLP. To express an APLP, transcriptional and translational signals recognizable by an appropriate host are necessary. The cloned APLP encoding sequences, obtained through the methods described above, and preferably in a double-stranded form, may be operably linked to sequences controlling transcriptional expression in an expression vector, and introduced into a host cell, either prokaryote or eukaryote, to produce recombinant APLP or a functional derivative thereof. Depending upon which strand of the APLP encoding sequence is operably linked to d e sequences controlling transcriptional expression, it is also possible to express APLP antisense RNA or a functional derivative thereof. Expression of the APLP in different hosts may result in different post- translational modifications which may alter the properties of the APLP. The present invention encompasses the expression of the APLP, or a functional derivative thereof, in eukaryotic cells, and especially mammalian, insect and yeast cells. Especially preferred eukaryotic hosts are mammalian cells either in vivo, in animals or in tissue culture. Mammalian cells provide post- translational modifications to recombinant APLP which include folding and/or glycosylation at sites similar or identical to that found for a native APLP. Most preferably, mammalian host cells include brain and neuroblastoma cells. A nucleic acid molecule, such as DNA, is said to be "capable of expressing" a polypeptide if it contains expression control sequences which contain transcriptional regulatory information and such sequences are "operably linked" to the nucleotide sequence which encodes die polypeptide. An operable linkage is a linkage in which a sequence is connected to a regulatory sequence (or sequences) in such a way as to place expression of the sequence under die influence or control of the regulatory sequence. Two DNA sequences (such as an APLP encoding sequence and a promoter region sequence linked to the 5 ' end of the encoding sequence) are said to be operably linked if induction of promoter function results in the transcription of the APLP encoding sequence mRNA and if die nature of the linkage between the two DNA sequences does not (1) result in d e introduction of a frame-shift mutation, (2) interfere with the ability of the expression regulatory sequences to direct the expression of the APLP mRNA, antisense RNA, or protein, or (3) interfere with the ability of the APLP template to be transcribed by the promoter region sequence. Thus, a promoter region would be operably linked to a DNA sequence if die promoter were capable of effecting transcription of that DNA sequence. The precise nature of the regulatory regions needed for gene expression may vary between species or cell types, but shall in general include, as necessary, 5 ' non-transcribing and 5 ' non-translating (non-coding) sequences involved wim initiation of transcription and translation respectively, such as the TATA box, capping sequence, CAAT sequence, and die like. Especially, such 5 ' non-transcribing control sequences will include a region which contains a promoter for transcriptional control of the operably linked gene. Expression of d e APLP in eukaryotic hosts requires the use of regulatory regions functional in such hosts, and preferably eukaryotic regulatory systems. A wide variety of transcriptional and translational regulatory sequences can be employed, depending upon me nature of the eukaryotic host. The transcriptional and translational regulatory signals can also be derived from me genomic sequences of viruses which infect eukaryotic cells, such as adenovirus, bovine papilloma virus, Simian virus, herpes virus, or the like. Preferably, these regulatory signals are associated widi a particular gene which is capable of a high level of expression in the host cell. In eukaryotes, where transcription is not linked to translation, such control regions may or may not provide an initiator methionine (AUG) codon, depending on whedier the cloned sequence contains such a methionine. Such regions will, in general, include a promoter region sufficient to direct the initiation of RNA synthesis in the host cell. Promoters from heterologous mammalian genes which encode mRNA product capable of translation are preferred, and especially, strong promoters such as the promoter for actin, collagen, myosin, etc., can be employed provided they also function as promoters in the host cell. Preferred eukaryotic promoters include those described above the promoter of the mouse metallothionein I gene (Hamer et al., J. Mol. Appl. Gen. 7:273-288 (1982)); the TK promoter of Herpes virus (McKnight, S. , Cell 37:355-365 (1982)); the SV40 early promoter (Benoist et al , Nature (London) 290:304-310 (1981)); in yeast, the yeast gal4 gene promoter (Johnston et al , Proc. Natl. Acad. Sci. USA 79:6971-6975 (1982); Silver et al. , Proc. Natl. Acad. Sci. USA 57:5951-5955 (1984)) or a glycolytic gene promoter may be used. As is widely known, translation of eukaryotic mRNA is initiated at die codon which encodes die first methionine. For this reason, it is preferable to ensure that the linkage between a eukaryotic promoter and a DNA sequence which encodes an APLP, or a functional derivative thereof, does not contain any intervening codons which are capable of encoding a memionine. The presence of such codons results either in a formation of a fusion protein (if the AUG codon is in d e same reading frame as the APLP encoding DNA sequence) or a frame-shift mutation (if the AUG codon is not in d e same reading frame as the APLP encoding sequence). If desired, a fusion product of an APLP may be constructed. For example, the sequence coding for APLP may be linked to a signal sequence which will allow secretion of d e protein from, or the compartmentalization of the protein in, a particular host. Such signal sequences may be designed widi or wi iout specific protease sites such that the signal peptide sequence is amenable to subsequent removal. Alternatively, the native signal sequence for diis protein may be used. Transcriptional initiation regulatory signals can be selected which allow for repression or activation, so that expression of the operably linked genes can be modulated. Of interest are regulatory signals which are temperature- sensitive so mat by varying the temperature, expression can be repressed or initiated, or are subject to chemical regulation, e.g., metabolite. Also of interest are constructs wherein APLP mRNA and antisense RNA are provided in a transcribable form, but with different promoters or other transcriptional regulatory elements such that induction of APLP mRNA expression is accompanied by repression of antisense RNA expression, and/or repression of APLP mRNA expression is accompanied by induction of antisense RNA expression. Translational signals are not necessary when it is desired to express APLP antisense RNA sequences. If desired, die non-transcribed and/or non-translated regions 3 ' to the sequence coding for APLP can be obtained by die above-described cloning metiiods. The 3 '-non-transcribed region may be retained for its transcriptional termination regulatory sequence elements; the 3-non-translated region may be retained for its translational termination regulatory sequence elements, or for those elements which direct polyadenylation in eukaryotic cells. Where me native expression control sequence signals do not function satisfactorily host cell, then sequences functional in the host cell may be substituted. -31- The vectors of the invention may further comprise other operably linked regulatory elements such as enhancer sequences, or DNA elements which confer tissue or cell-type specific expression on an operably linked gene. To transform a mammalian cell with the DNA constructs of die invention many vector systems are available, depending upon whemer it is desired to insert an aAPLP DNA construct into the host cell chromosomal DNA, or to allow it to exist in an extrachromosomal form. If the APLP DNA encoding sequence and an operably linked promoter is introduced into a recipient eukaryotic cell as a non-replicating DNA (or RNA) molecule, which may either be a linear molecule or a closed covalent circular molecule which is incapable of autonomous replication, the expression of die APLP may occur through the transient expression of the introduced sequence. Genetically stable transformants may be constructed with vector systems, or transformation systems, whereby APLP DNA is integrated into die host chromosome. Such integration may occur de novo within the cell or, in a most preferred embodiment, be assisted by transformation with a vector which functionally inserts itself into the host chromosome, for example, wi retroviral vectors, transposons or other DNA elements which promote integration of DNA sequences in chromosomes. A vector is employed which is capable of integrating the desired gene sequences into a mammalian host cell chromosome. Cells which have stably integrated the introduced DNA into dieir chromosomes are selected by also introducing one or more markers which allow for selection of host cells which contain the expression vector in the chromosome, for example, me marker may provide biocide resistance, e.g., resistance to antibiotics, or heavy metals, such as copper, or the like. The selectable marker gene can either be directly linked to d e DNA gene sequences to be expressed, or introduced into die same cell by co-transfection. In another embodiment, me introduced sequence is incorporated into a plasmid or viral vector capable of autonomous replication in me recipient host. Any of a wide variety of vectors may be employed for this purpose, as oudined below. Factors of importance in selecting a particular plasmid or viral vector include: d e ease widi which recipient cells that contain the vector may be recognized and selected from those recipient cells which do not contain the vector; the number of copies of the vector which are desired in a particular host; and whedier it is desirable to be able to "shuttle" die vector between host cells of different species. Preferred eukaryotic plasmids include diose derived from the bovine papilloma virus, vaccinia virus, SV40, and, in yeast, plasmids containing the 2-micron circle, etc., or their derivatives. Such plasmids are well known in the art (Botstein et al. , Miami Wntr. Symp. 79:265-274 (1982); Broach, J.R., In: The Molecular Biology of the Yeast Saccharomyces: Life Cycle and Inheritance, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, pp. 445-470 (1981); Broach, J.R., Cell 25:203-204 (1982); Bollon et al. , J. Clin. Hematol. Oncol. 70:39-48 (1980); Maniatis, T., In: Cell Biology: A Comprehensive Treatise, Vol. 3, "Gene Expression," Academic Press, NY, pp. 563-608 (1980)), and are commercially available. For example, mammalian expression vector systems which utilize die MSV-LTR promoter to drive expression of die cloned gene, and in which it is possible to cotransfect with a helper virus to amplify plasmid copy number, and integrate the plasmid into die chromosomes of host cells have been described (Perkins et al. , Mol. Cell Biol. 3:1123 (1983); Clontech, Palo Alto, California). Once the vector or DNA sequence containing the construct(s) is prepared for expression, die DNA construct(s) is introduced into an appropriate host cell by any of a variety of suitable means, including transfection. After die introduction of die vector, recipient cells are grown in a selective medium, which selects for the growdi of vector-containing cells. Expression of the cloned gene sequence(s) results in the production of an APLP, or in the production of a fragment of this protein. This expression can take place in a continuous manner in the transformed cells, or in a controlled manner, for example, expression which follows induction of differentiation of die transformed cells (for example, by administration of bromodeoxyuracil to neuroblastoma cells or the like). The expressed protein is isolated and purified in accordance widi conventional conditions, such as extraction, precipitation, chromatography, affinity chromatography, electrophoresis, or the like. The APLP DNA encoding sequences, obtained through the methods above, will provide sequences which, by definition, encode an APLP and which may then be used to obtain APLP antisense RNA genetic sequences as die antisense RNA sequence will be that sequence found on die opposite strand of the strand transcribing the peptide core's mRNA. The antisense DNA strand may also be operably linked to a promoter in an expression vector such that transformation with this vector results in a host capable of expression of an APLP antisense RNA in the transformed cell. Antisense RNA and its expression may be used to interact widi an endogenous APLP DNA or RNA in a manner which inhibits or represses transcription or translation of an APLP gene in a highly specific manner. Use of antisense RNA probes to block gene expression is discussed in Lichtenstein, C, Nature 333:801-802 (1988). For example, such probes can be used to block me expression of an APLP when the expression is aberrant. Having generally described die invention, the same will be more readily understood dirough reference to the following examples which are provided by way of illustration and are not intended to be limiting. Experimental Example I Materials and Methods Cell culture and protein extracts Human neuroglioma H4 cells were maintained in DMEM medium supplemented widi 10% fetal calf serum, 1 % L-glutamine and 1 % penicillin- streptomycin at 37 °C in a humidified atmosphere containing 5% CO 2 /95% O 2 . Nuclear extracts were prepared according to Miner et al. (Miner, L. L. et al., J. Neurosci. Res. 33:10-18 (1992)). Final protein concentration varied between 0.5 and 2 mg/ml. Preparation of DNA fragments and plasmid constructs Two complementary oligonucleotides were synthesized for each DNA fragment employed in gel mobility shift assays. The oligonucleotides were annealed and end-labeled widi [α- 32 P]dCTP by fill-in reaction. The sequence of die DK-1 fragment extends from -30 to -58 from die primary transcriptional site of the human APP promoter (Salbaum, J. M. et al, EMBO J. 7:2807-2813 (1988)) and contains the AP-l/AP-4 element (underlined): 5'GGGCCGGAI£ΔQCIQΔCI£GCCTGGCTCT3' (SEQ ID NO:2). The randomized versions of the DK-1 fragment contain selectively randomized sequences, underlined as follows: 5'Random: 5'QCIIΔςiQTCAGCTGACTCGCCTGGCTCT3' (SEQ ID NO:4); AP-l/AP- 4 Random: 5 'GGGCCGGAATCGTGCTGTCGCCTGGCTCT3 ' (SEQ ID NO:5); 3' Random: 5OGGCCGGATCAGCTGACG-ΔT^CCIQICCQ3' (SEQ ID NO:6). The AP-1 fragment contains a consensus sequence for the c-fos/c- jun transcription factors (Mermod, N. et al , Nature 332:557-561 (1988)) (underlined): 5 'GATCCAGCTGACTC ATC ACTAG3 ' (SEQ ID NO:7). The consensus sequence for die AP-4 transcription factor (Hu, Y.-F. et al. , Genes Dev. 4:1741-1752 (1990)) is underlined in the following sequence: 5OATCACCΔQCIQTGGAATGTGTGTGATC (SEQ ID NO:8). Finally, the core sequence of die USF binding site (Gregor, P. D. et al , Genes Dev. 4:1730-1740 (1990)) is inserted into the USF fragment: 5OGGCCGGATCΔCGIGACTCGCCTGGCTCT3' (SEQ ID NO:9). The hAPP-luciferase construct was prepared by inserting a 2.9 kb EcoRII/BamHI APP promoter fragment (Salbaum et al, EMBO J. 7:3807 (1988)) into the Smal site of die luciferase expression vector pxP2 (Nordeen, S.K. Biotechniques 6:454 (1988)). The pMLΔ53(C 2 AT) plasmid was constructed as described (Roy et al, Nature 354: 245 (1991)). The pCMV- USF construct was prepared by inserting the gene encoding die 43 kDa USF protein into the pCMV vector. The pCMV-aplpl and pCMV-aplp2 constructs were prepared by inserting the gene encoding the aplpl and aplp2 proteins (described below) into die pCMV vector. Electrophoretic mobility shift assay (EMSA) EMSA was carried out as in Miner et al. (Miner, L. L. et al. , J. Neurosci. Res. 33:10-18 (1992)). 1 ng of DNA fragment was incubated with 1 μg nuclear extract for 20 min at 20°C, unless otherwise indicated. Quantities of rUSF (Pognonec, P. et al , Mol. Cell Biol 77:5125-5136 (1991)) are indicated in die figure legends. After incubation, the binding mixture was resolved on 6% polyacrylamide gels containing 4% glycerol in 25 mM Tris- borate (pH 8.3), 0.5 mM EDTA. Gels were dried and exposed to Kodak X- OMAT AR film with intensifying screen. -36- Western analysis and antibodies Protein extracts were size-fractionated on 8% polyacrylamide gels according to Laemmli (Laemmli, U. K. Nature 227:680-685 (1970)) and were transferred to Immobilon P (Millipore) electrophoretically at 100 V in 20 mM Tris (pH 7.4), 150 mM glycine and 20% methanol for 4 hr at 4°C. Membranes were blocked 10 mM Tris (pH 8.0), 150 mM NaCl, 0.05% Tween 20, 3% BSA, 0.05% NaN 3 for 1 hr at 20°C. After incubation with the USF antiserum (Kaulen, H. et al. , Mol. Cell. Biol. 77:412-424 (1991)), the membranes were immunostained using a light-emitting luminol/horseradish peroxidase system (ECL Western blotting; Amersham), according to d e manufacturer's protocol. The c-fos antiserum (Oncogene Products, Manhasset, NY) was prepared against the DNA-binding epitope of die c-fos protein. In vitro transcription and primer extension In vitro transcription was performed according to Dignam, J.D. et al, Nucleic Acids. Res. 77: 1475-1489 (1983), modified as recommended by the manufacturer (Promega Corp.). Primer extension was carried out on die transcription product as described in Martinez, E. et al , EMBO J. 13:3115- 3126 (1994). The primer specific for the hAPP-luciferase construct corresponds to part of the multiple cloning site between me promoter and die reporter gene widi die following sequence: 5'-GCTCAGATCTCGAGCTCGGTAC-3' (SEQ ID NO:3). The primer for the MLΔ53 plasmid encompasses 19 bp of me G-less cassette: 5'-GGAAATATAGAAGAAGGAG-3' (SEQ ID NO:10). RNA and end-labeled primers were hybridized in 50% formamide, 10 mM Tris (pH 7.5), 250 mM KCl and 1 mM EDTA for 10 min at 65°C and overnight at 42°C. Sequencing reaction was performed using die APP primer extension 6/15265 -37- oligonucleotide according to die directions suggested by die manufacturer (United States Biochemical Corp.). Results DNA-Protein Binding at the AP-l/AP-4 Site Using H4 Cell Nuclear Extracts To examine whether the AP-l/AP-4 element binds to transcription factors present in the nucleus of H4 neuroglioma cells, a 29 bp 32 P-labeled doublestranded DNA fragment (DK-1, see FIGURE 1A (SEQ ID NO:2)) containing the AP-l/AP-4 site was incubated in the presence of H4 nuclear extracts and separated on a non-denaturing polyacrylamide gel (electrophoretic mobility shift assay, EMSA). One discrete band was observed (FIGURE IB). The specificity of the binding was assessed by die addition of increasing amounts of cold fragment, which resulted in a very rapid decrease of the binding on die labeled fragment. To identify die region within die 29 bp DK-1 fragment that is responsible for die binding, diree differentially mutated double-stranded DNA fragments were synmesized for use in competition assays. The first of these contained a random sequence in place of the region upstream of the AP-l/AP-4 site, the second contained a random sequence in place of the AP-l/AP-4 element itself, and die diird contained a random sequence in place of die region downstream from the AP-l/AP-4. Binding to die labeled fragment was significantly diminished only in me presence of me two competitor fragments that still contained an intact AP-l/AP-4 site, while the competitor fragment that contained a random sequence in place of the AP-l/AP-4 site did not effectively compete for binding. These results indicate diat die H4 nuclear extract contains a factor that specifically binds to d e AP-l/AP-4 element of the APP promoter. DNA-Protein Interaction at the AP-l/AP-4 Site Involves a Factor Different from the c-Fos/c-Jun or AP-4 Factors The c-fos/c-jun complex is known to interact with the AP-1 sequence alone. Therefore, to determine whedier the protein binding to die combined AP-l/AP-4 site was either die c-fos/c-jun complex or the AP-4 factor, EMSA was performed in die presence of DNA fragments containing consensus sequences for the AP-1 and die AP-4 elements and in die presence of a monoclonal antibody directed against the DNA binding site of c-fos (FIGURE 2). None of these agents affected die binding to die AP-l/AP-4 site in our system. Next, specific binding to the AP-1 consensus sequence was assessed by labeling d is fragment and incubating it widi die H4 nuclear extract. The resulting DNA-protein complex migrated more slowly man that observed using the AP-l/AP-4 element (see FIGURE 4B), was specifically diminished in die presence of increasing amounts of c-fos antibody, and was not affected by die presence of a 50-fold excess of DK-1 fragment (data not shown). Collectively, diese data indicate that die binding at die AP-l/AP-4 site does not involve die c-fos/c-jun complex or die AP-4 factor. The Factor Binding the AP-l/AP-4 Site Also Interacts with the USF Binding Site A double-stranded DNA fragment incorporating me core sequence of die consensus element for USF, CACGTG, was labeled and incubated widi the H4 nuclear extract. The resulting complex exhibited a very similar pattern of migration to diat obtained with die AP-l/AP-4 site in the core position, and was specifically and rapidly diminished following the addition of cold DK-1 fragment to the incubation mixture (FIGURE 3). Conversely, the formation of the complex using the AP-l/AP-4 element was also rapidly abolished in die presence of the cold USF fragment. These experiments indicate that die DNA- protein complexes formed at die AP-l/AP-4 and USF sites include eidier the same or related factors. Antiserum Raised Against USF Recognizes the Complex on the AP-l/AP-4 S e We next investigated whedier USF is a component of the DNA-protein complex formed widi die AP-l/AP-4 site. For diis purpose we first tested for the presence of USF in the H4 nuclear extract with a polyclonal antiserum raised against the 43 KDa form of USF and demonstrated the presence of a single band at around 43 kDa, which comigrates with die recombinant USF protein (FIGURE 4A). The same anti-USF antiserum also had the ability to significandy modify die migration rate of the complex formed on die AP-l/AP- 4 site (FIGURE 4B). It caused a marked decrease in die amount of normal DNA-protein complex formation, and resulted in die formation of a new, more slowly migrating complex. This complex is the result of an interaction of die USF antibody widi die DNA-protein complex and dius constitutes a "supershift. " Increased amounts of anti-USF antiserum resulted in die elimination of die slower migrating band. The specificity of the complex formation was confirmed by showing diat die anti-USF antiserum did not affect the complex formation on the AP-1 site alone. Control antibodies directed against another protein, APP 751 , did not affect the complex formation on the AP-l/AP-4 site (data not shown). Taken together, these data indicate diat die factor present in the DNA-protein complex formed widi me AP-l/AP-4 element is antigenically related to USF. Recombinant 43 KDa USF Binds to the AP-l/AP-4 Element In order to demonstrate a direct interaction between the AP-l/AP-4 site and USF, we employed the 43 kDa form of USF expressed in, and purified from, bacteria. This recombinant USF (rUSF) polypeptide has been shown to bind to its consensus sequence in EMSA, exhibiting a pattern very similar to that obtained widi various nuclear extracts (Pognonec, P. et al. , Mol. Cell Biol 77:5125-5136 (1991)). Incubation of the DK-1 fragment with small amounts (350 pg) of rUSF resulted in the formation of a visible complex, exhibiting the same migration pattern obtained widi die H4 nuclear extract (FIGURE 5). The amount of complex formed decreased with increasing amounts of cold fragment. The specificity of e binding of rUSF to the AP-l/AP-4 site was demonstrated dirough competition experiments using non-radioactive DK-1 fragment in which the AP-l/AP-4 site was replaced by a randomized sequence. The addition of cold DK-1 fragment containing randomized AP-l/AP-4 site failed to diminish DNA-protein formation on the labeled DK-1 fragment (data not shown). These results indicate mat rUSF binds specifically to the AP- l/AP-4 site, and strongly suggest that die USF protein present in the H4 cell nuclear extract is partly or fully responsible for the formation of the DNA- protein complex on the AP-l/AP-4 site present in the APP promoter. The Presence of USF Maintains Elevated Levels of Constitutive APP Expression To test the ability of USF to modulate transcription from the APP promoter, cell-free in vitro transcription reactions were performed using HeLa cell nuclear extracts in the presence of increasing amounts of the DNA fragment containing the consensus element for USF. The presence of USF in HeLa cell nuclear extracts has previously been characterized (Pognonec, P. et al , Mol. Cell. Biol. 77:5125-5136 (1991)) and confirmed in diis study by Western Blot analysis (data not shown). USF levels are higher in HeLa nuclear extracts than in die H4 neuroglioma extracts, as indicated by increased binding to the DK 1 fragment (FIGURE 6A). In the cell-free transcription assays, the correct transcription initiation site was demonstrated in die hAPP-luciferase construct by primer extension and by direct sequencing widi a primer located downstream from the BamHI site at position + 105. Additional transcription initiation sites were also found (FIGURE 6B), and correspond to tiiose reported by La Fauci, G. et al., Biochem. Biophys. Res. Commun. 759:297-304 (1989). Two constructs, one containing the APP promoter and one containing 53 bp of the adenovirus major late promoter lacking a USF site (Roy, A.L. et al., Nature 354:245-248 (1991), were used in the same in vitro transcription reaction. FIGURE 6C shows that the addition of a 20-fold molar excess of the DNA fragment containing the USF consensus site significantly decreases transcription from the APP promoter. Conversely, the presence of the DNA fragment in which die AP-l/AP-4 site was replaced by a random sequence does not decrease transcription at the same molar concentration. The control construct indicated similar levels of transcriptional efficiency in the reactions. These data indicate diat the presence of USF in me nuclear extract is important for maintaining the basal level of transcription from the human APP promoter. Recombinant 43 KDa USF Activates Transcription from the APP Promoter To examine whether USF can increase transcription from the APP promoter above basal levels, 100 ng of rUSF were added to die in vitro transcription reactions, followed by primer extension analysis (FIGURE 7). The amount of transcription product doubled under tiiese conditions. 50 ng of rUSF were sufficient to yield a similar increase (data not shown). The high levels of USF already present in the HeLa nuclear extracts may serve to mask a more robust increase in transcription levels following the addition of rUSF. These results suggest that USF participates in the activation of transcription from the APP promoter, and diat levels of USF in die nucleus strongly correlate widi die amount of APP mRNA synthesized. Recombinant 43 KDa USF and the APP/APLP family modulate transcription from the APP promoter H4 neuroglioma cells were cotransfected with three constructs: die APP promoter-luciferase gene construct, the USF expression vector (pCMV-USF) or its control plasmid, and die APP/APLP family expression vectors (pCMV- aplpl, pCMV-aplp2, and pCMV-APP) or their control plasmid. The results were assayed in terms of relative luciferase units and showed diat expressing USF alone in the neuroglioma cells activated transcription from the APP promoter about 5-fold as compared to die control. However, when eidier APLPl, APLP2, or APP was also expressed in the cells, in each instance, USF activation of transcription from die APP promoter decreased to diat of about 2- fold as compared to die control. These results are shown graphically in FIGURE 8. Example 2 Materials and Methods Neuroblastoma NB2A cells were maintained as previously described (Magendantz et al., Proc. Natl. Acad. Sci. USA 52:6581-6585 (1985)). Radionucleotides were obtained from New England Nuclear and Amersham. Restriction enzymes were obtained from New England Biolabs and PCR reagents from Perkin-Elmer. Screening of λgtll Libraries. General techniques for preparing and screening libraries are disclosed in Maniatis et al , Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, 2d ed. (1989). Three different libraries were used to obtain mouse brain cDNA clones in λgtl l. A random primed and an oligo-dT primed library were obtained from Clontech. An oligo-dT primed library was obtained from Stratagene. Libraries were screened by hybridization to nitrocellulose (BA85, Schleicher and Schuell) or Nylon (Hybond-N, Amersham) according to standard procedures using cDNA that was labeled by random priming (Feinberg et al, Anal. Biochem. 132:6-13 (1983)). Positive clones were sized by PCR amplification of the λgtll insert using primers 1218 and 1222 from New England Biolabs. Recombinant DNA Techniques DNA fragments were subcloned into pBluescript (Stratagene) or Ml 3 (New England Biolabs) vectors and botii strands were sequenced widi Sequenase (U.S. Biochemical) according to die manufacturer's instructions. Sequence analyses were done using the UWGCG programs at the Whitaker College Computing Facility at MIT. RACE Procedure for Obtaining 5' cDNA Extensions The RACE (Rapid Amplification of cDNA Ends) procedure diat was used is a combination of me methods of Frohman et al. (Frohman et al. , Proc. Natl. Acad. Sci. USA 55:8998-9002 (1988)) and Ohara et al. (Ohara et al , Proc. Natl. Acad. Sci. USA 56:5673-5677 (1989)). For the RACE procedure, die primers were the complements of nucleotides 699-719 and 672-692 of die sequence presented in FIGURE 10 (SEQ ID NO: 15). RACE products were subcloned into pBluescript, screened by hybridization to die 5' 120 bp EcσRI- Pstl fragment of 69A and positive clones were sequenced. RNA Analysis Poly A + RNA was prepared as in Badley et al. (Badley et al., BioTechniques 6:114-116 (1988)), using oligo-dT beads (Collaborative Research). For Northern blot analysis the RNA was separated on an agarose gel containing formaldehyde, transferred to nylon (BioTrace, Gelman Sciences) according to standard methods (Sambrook et al , Molecular Cloning (A Laboratory Manual), Cold Spring Harbor Laboratory, (1989)), and crosslinked to die nylon using a UV Crosslinker (Stratagene). The blots were hybridized and washed according to the metiiod of Church and Gilbert (Church et al, J. Cell Biol. 107:1765-1772 (1988)). The molecular weight of the transcripts was determined by using RNA molecular weight markers. Production Antisera to an APLP Peptide. A peptide with the sequence QQLRELQRH (SEQ ID NO:l) was obtained from the Biopolymers laboratory of the Howard Hughes Medical Institute and Center for Cancer Research at MIT. 20 mg of the peptide was conjugated to KLH essentially as described in Marcantonio, C.G., and Hynes, R.O., J. Cell Biol. 107:1765-1772 (1988), and the immunization of four New Band white rabbits was carried out as described in Schatz et al. (Schatz et al, Mol. Cell Biol. 7:3799-3805 (1987)). Protein Preparation Protein from neuroblastoma cells was isolated by rinsing die cells widi PBS followed by lysing die cells in SDS sample buffer and boiling. Protein from mouse brain was isolated by homogenizing one brain in 1ml of RIPA buffer (50 nM Tris pH 7.4, 150 mM NaCl, 5 mM EDTA, 1% Triton X-100, 1 % Na deoxycholate, 0.1 % SDS and protease inhibitors). The homogenate was spun in an Eppendorf centrifuge for 30 minutes at 4°C, combined widi SDS sample buffer and boiled. β-galactosidase Fusion Protein Preparation A β-galactosidase-APLPl fusion protein was constructed using standard techniques (Sambrook et al. , Molecular Cloning (A Laboratory Manual), Cold Spring Harbor Laboratory, (1989)). An EcόBl-EcόBl fragment from a λgtll clone containing 666 nucleotides of die 3' coding portion (nucleotides 1380- 2046 of FIGURE 10 (SEQ ID NO: 15)), and 258 nucleotides of the untranslated region of the APLP (nucleotides 2047-2305 of FIGURE 10 (SEQ ID NO: 15)) was ligated into the EcoRI site of a pUΕX5 vector. To prepare total protein lysates from bacteria containing these plasmids, an overnight culture, diluted 1:10 into L-broth plus ampicillin, was grown for 1.5-2 hours at 300, dien induced at 42° (or left at 30° for uninduced samples) for 2.5-3 hours. The bacteria were then spun down, resuspended in 50% SDS sample buffer containing protease inhibitors and sonicated to shear chromosomal DNA. Western Blot Analysis Protein samples were subjected to polyacrylamide gel electrophoresis, transferred to nitrocellulose and probed widi rabbit antibodies and 125 I-labelled protein A essentially as described in Birgbauer (Birgbauer et al, J. Cell Biol. 709:1609-1620 (1989)). Immunofluorescence Neuroblastoma cells were plated onto glass coverslips approximately forty eight hours before fixation. Twenty four hours before fixation, the concentration of fetal calf serum in the medium of neuroblastoma cells was changed from 10% to 0.1 % to induce neurite extension. Twenty minutes before fixation, concanavalin A was added to 20 mg/ml to encourage cell adhesion to die coverslips. Cells were fixed in 3.7% formaldehyde/PBS, permeabilized in acetone and blocked for 30 minutes at 37° in PBS containing 1 % calf serum. Primary antibody was diluted into the blocking buffer, applied to die cells for 30 minutes and visualized widi FITC conjugated goat anti-rabbit antibody. Cells were observed and photographed using a Zeiss Axioplan microscope. For the peptide competition experiment, the peptide was preincubated widi die primary antibody in blocking buffer for 30 minutes before adding it to the cells (Donaldson et al., J. Cell Biol 777:2295-2306 (1990) and Moremen et al, J. Biol. Chem. 260:6654-6662 (1985)). Results Identification and Cloning of APLPl In a screen for cDNA clones encoding a microtubule-associated protein (MAP), a clone was isolated from a mouse brain cDNA library (Stratagene) which was found to have an open reading frame (ORF) homologous to that of APP. The probe that was used to screen was an antibody elicited against MAP. APLPl is not related to any known MAP. The cDNA clone in which the APP homology was originally identified contained a portion of die C-terminal coding sequence as well as a portion of the 3' untranslated region. To extend die APLPl ORF in the 5' direction, probes were used from die 5 '-most regions of available cDNA clones to screen two Clontech λgtll libraries. Repetitive screens using progressively more upstream probes resulted in die isolation of a 1.8 kb cDNA clone, 69A (FIGURE 9), whose 5' terminus has an EcoRI site that is present in the coding sequence of APLPl and is die result of an EcoRI site that escaped methylation during construction of the cDNA library. Aldiough screening of the cDNA libraries widi probes derived from the 5' end of 69A failed to identify any more APLPl clones, the use of a variation of the RACE procedure developed by Frohman et al. (Frohman et al. , Proc. Natl Acad. Sci. USA 55:8998-9002 (1988)) and by Ohara et al. (Ohara et al, Proc. Natl Acad. Sci. USA 56:5673-5677 (1989)) did enable die isolation of several independent, overlapping cDNA clones that extend me APP homology past the 5' EcoRI site of 69A. The longest clone, J (FIGURE 1), did not contain an initiator methionine. The sequence information obtained via die RACE procedure was used to create PCR primers and amplify the 5 '-most 100 base pairs encoded by clone J. This PCR product was used as a probe in a screen of a Stratagene mouse brain cDNA library that successfully identified a number of full length APLPl clones. Two of these were sequenced to obtain the final 313 5' nucleotides as well as d e polyadenylation signal and die poly A tail of the APLPl cDNA. The predicted initiator methionine is in agreement with die eukaryotic consensus initiation sequence (Kozak, M., Nucl. Acids Res. 72:857-872 (1984). APLPl is Related to APP The 2361 nucleotides of die cDNA sequence encode an open reading frame of 653 amino acids as is shown in FIGURE 10 (SEQ ID NO: 16). The protein is predicted to have a short intracellular C-terminus of 46 amino acids, a membrane spanning domain of 23 amino acids, and a larger extracellular N- terminus. The predicted amino acid sequence and die overall structure of APLPl is similar to those of APP, which resembles an integral membrane protein (Kang et al, Nature 325:733-736 (1987)). The alignment of die two amino acid sequences that is shown In FIGURE 11 (SEQ ID NOS:17-18) reveals that overall APLP is 42% identical and 64% similar to APP. APLP1 is a Member of a Family of APP-like Proteins The identities between APLPl and APP are concentrated in three distinct regions (FIGURE 12 (SEQ ID NOS: 19-28)), where the proteins are 47, 54 and 56% identical and 67, 73 and 74% similar. These same three regions have been shown previously to be shared between APP and a Drosophila APP- like protein (Drosophila APPL), and have been termed die extracellular I (El), extracellular II (EII) and cytoplasmic (C) domains by these investigators (Rosen et al, Proc. Natl. Acad. Sci. USA 56:2478-2482 (1989)). The cytoplasmic domain homology is also present in a partial cDNA clone that has been isolated from a rat testis library (Yan et al. , Proc. Natl. Acad. Sci. USA 57:2405-2408 (1990)). Only APP contains the βA4 sequence diat is found in amyloid plaques. FIGURE 12 (SEQ ID NOS: 19-28) shows die domain alignment of die four proteins mentioned above. A similar alignment has been shown for die relationship between die Drosophila APPL1 and APP (Rosen et al, Proc. Natl. Acad. Sci. USA 56:2478-2482 (1989)). The testis cDNA is included only in the C domain comparison since only this portion of the predicted amino acid sequence is known (Yan et al. , Proc. Natl Acad. Sci. USA 57:2405-2408 (1990)). El begins at amino acid 21 in me APLPl open reading frame and spans 136 amino acids. Overall, 102 of tiiese 136 amino acids (75%) are eitiier identical to amino acids in me respective positions of APP or Drosophila APPL, or they are the same in all tiiree proteins. The most striking conservation widiin this region is that of 12 cysteine residues in all three of the sequences. There are also two regions of amino acids diat are particularly well conserved (underlined in FIGURE 12 (SEQ ID NOS: 19-28)), as is an unusually acidic region composed of glutamic and/or aspartic acids diat spans amino acids 237-271 in the APLPl sequence (FIGURE 10 (SEQ ID NO: 16)). EII spans 130 amino acids in die mouse APLPl sequence. 93 of die 130 APLPl amino acids (71 %) are identical to eitiier one or both of their counterparts in APP or the Drosophila APPL sequences. This region also contains conserved N-glycosylation site in all 3 proteins. The third domain encompasses the C-terminal cytoplasmic region of all of the proteins, including die predicted amino acid sequence of d e rat testis cDNA. The conservation of amino acids among the members of the APP-like family within this domain is particularly strong. Although die four proteins do not share homology within the predicted transmembrane domains (FIGURE 11 (SEQ ID NOS: 17-18); Rosen et al, Proc. Natl. Acad. Sci. USA 56:2478-2482 (1989); Yan et al , Proc. Natl. Acad. Sci. USA 57:2405-2408 (1990)), all of them do contain a 3-4 amino acid span of charged residues (arginine/lysine) at the cytoplasmic face of the membrane (FIGURE 12 (SEQ ID NOS: 19-28)). This characteristic is often seen at the membrane-cytoplasmic junction of odier proteins, and has been hypotiiesized to allow for an interaction with phospholipids in die membrane, or to provide a stop transfer signal for membrane bound proteins (Blobel, G., Proc. Natl. Acad. Sci. USA 77:1796- 1500 (1980)). Northern Blot Analysis FIGURE 13 shows autoradiographs of Northern blots containing poly A + RNA from mouse brain and neuroblastoma cells mat were probed with DNA corresponding to nucleotides 1791-2305 of FIGURE 10 (SEQ ID NO: 15). These blots reveal that in mouse brain and neuroblastoma cells mere are two messages of approximately 2.4 and 1.6 kb diat hybridize to diis probe. The larger message appears to be present in relatively greater abundance tiian the smaller message. Because of its size, it is clear that die cDNA diat corresponds to die 2.4 kb message, although botii messages are consistently seen in Northerns that are probed and washed under stringent conditions. The mouse APLPl cDNA docs not hybridize to the 3.2 and 3.4 kb APP messages under die conditions used (see Materials and Metiiods; Kang, J. et al. , Nature 325:733-736 (1987)). Generation of Antibodies Against an APLPl Peptide. In order to further characterize the protein encoded by die APLPl cDNA, antibodies were raised to a syntiietic peptide which corresponds to a unique sequence of mouse APLPl . The peptide d at was used as antigen corresponds to a 9 amino acid segment located near the C-terminus of the .APLPl protein (QQLRELQRH) (SEQ ID NO: l), a region where the four proteins are not homologous. Four rabbits were injected wim the peptide as described in Materials and Metiiods. Two of die four rabbits (301 and 302) produced sera that strongly recognize a 65 kDa mouse brain protein that is not recognized by die appropriate preimmune sera (FIGURE 14A). A smaller protein of approximately 33 kDa that is recognized by antiserum 301 may be a proteolytic degradation product of die larger protein. In FIGURE 14A, the specificity of the interaction of die antibody widi tiiese proteins is demonstrated by die ability to block the binding of antibody 301 to die proteins by preabsorbing widi die original peptide (lanes 2-5); an irrelevant peptide has no effect on die interaction of the antibody widi either the 65 kDa or 33 kDa protein (lane 6). Antiserum 301 also recognizes a 65 kDa protein present in neuroblastoma cell extracts that is not recognized by preimmune serum (FIGURE 14B). To further confirm e specificity of the 301 antiserum, we determined whedier the antiserum would recognize a β-galactosidase fusion protein containing die 222 carboxy-terminal amino acids encoded by die APLPl cDNA. FIGURE 14C shows a Western blot of bacterially produced proteins that were probed widi antiserum 301. As can be seen in lane 6 of this figure, antisera 301 does specifically interact with die β-galactidase- APLPl fusion protein. There are a number of antibodies diat have been generated against the C-terminus of APP. Because the identity between APP and die mouse APLPl in this region is particularly strong, some of these antisera would also be likely to interact with die mouse APLPl. One of these antisera, R37 (Kang et al, Nature 325:733-736 (1987); and Ishii et al, Neuropatolo. andAppl. Neurobiol 75:135-147 (1989)), is directed against the carboxy-terminal 15 amino acids of aAPP, a region where die two proteins are particularly similar (see FIGURE 12 (SEQ ID NOS: 19-28)). R37 does recognize die β-galactosidase-APLPl fusion protein and a 65 kDa mouse brain protein that comigrates widi die 65 kDa protein recognized by antiserum 301 (data not shown). The 15 amino acid sequence used to raise the anti-APP antibody does not overlap the 9 amino acids used to generate antiserum 301. These data suggest d at the 65 kDa protein contains two epitopes in common with die APLPl fusion protein. Antibodies can be made which recognize only APLPl . Anti-APLPl Antisera Recognizes a Protein in the Golgi. The subcellular localization of die protein recognized by antiserum 301 was assayed by immunofluorescence. When neuroblastoma cells are stained widi 301, the pattern diat is observed is a reticular staining near the nucleus (FIGURE 15A,B). Because of 3-dimensional nature of the staining, and die round shape of die cells, die image seen in any one plane of focus appears punctate rather tiian reticular. An identical pattern is seen with antiserum 302 (data not shown). The pattern itself is reminiscent of Golgi staining, and when tiiese cells are stained with an antibody to a known Golgi enzyme, mannosidase II, a pattern much like that seen widi antiserum 301 is observed (FIGURE 15C). The inclusion of die original peptide in die antibody incubation inhibited the 301 staining (FIGURE 15D). Staining was not seen when preimmune serum was used (FIGURE 15E). Discussion The present APLPl cDNA sequence encodes a new member of the APP-like family. The mouse homologue of the human amyloid precursor protein has been cloned previously and is 96.8% identical to die human sequence at the amino acid level (Yamada et al, Biochem. Biophys. Res. Comm. 755:906-912 (1987)). Thus, the present APLPl cDNA, which is 42% identical to die amyloid precursor protein at the amino acid level, is not the mouse homologue of APP and is a distinct, yet related protein. The two sequences share three domains of homology. The amino acid conservation within these domains include 12 cysteines, an unusually acidic region, a potential N-glycosylation site, a hydrophobic membrane spanning region, and several specific blocks of exact identity. It is clear that the mouse APLP, the Drosophila APPL, the rat testis protein and APP comprise a family of proteins. The extensive conservation of amino acid identity as well as botii the overall and specific domain structure witiiin this family of proteins suggests that tiiese proteins share a common function. There are two potentially interesting observations that can be made concerning the strict conservation of the 7-amino acid sequence located within die cytoplasmic tail of the proteins in the APP-like family (see underlined sequence in the appropriate portion of FIGURE 12 (SEQ IS NOS: 19-28)). There is a potential tyrosine phosphorylation site present 8-9 amino acids from the carboxy terminus of all four sequences (Tamkun et al, Cell 46:271-282 (1986)). APP can be phosphorylated when introduced into transformed embryonic kidney cells (Oltersdorf et al, J. Biol. Chem. 265:4492-4497 (1990)) and a peptide containing a portion of die cytoplasmic domain can be phosphorylated on serine and direonine residues in vitro (Gandy et al. , Proc. Natl. Acad. Sci. USA 55:6218-6221 (1988)), but tyrosine phosphorylation has not yet been demonstrated. Agents that are known to regulate protein phosphorylation appear to affect the rate of proteolytic processing of mature forms of APP (Buxbaum et al , Proc. Natl. Acad. Sci. USA 57:6003-6006 (1990)), suggesting that abnormal protein phosphorylation may be involved in the production of βA4. The sequence surrounding this tyrosine also shares homology with die only tyrosine in the o-helical domain that is conserved between several classes of intermediate filaments (Lendahl et al, Cell 60:585- 595 (1990)). The conservation of this potentially phosphorylated tyrosine is intriguing in light of me role that tyrosine phosphorylation is known to play in the regulation of cell growth and differentiation. The same tyrosine is part of the tetrameric sequence NPxY diat is believed to be required for the ligand-independent, coated pit-mediated internalization of the low density lipoprotein receptor (Chen, et al, J. Biol. Chem. 265:3116-3123 (1990)). The NPxY sequence is present in the cytoplasmic tails of at least 16 other cell surface receptor molecules -including die β-integrin receptor and members of the EGF receptor family (Chen, et al, J. Biol. Chem. 265:3116-3123 (1990)). The APLPl cDNA that has been isolated shares at least one epitope with a 65 kDa protein that is present in mouse brain homogenates and neuroblastoma cell extracts, and it shares an epitope with a protein diat localizes to the Golgi in neuroblastoma cells. An antiserum that recognizes die Drosophila APPL protein also recognizes a protein in the Golgi (Luo et al, J. Neurosci 70:3849-3861 (1990)). In addition, antibodies to the APP give a perinuclear staining pattern suggestive of either Golgi or ER localization when used for immunofluorescence on muscle fibers (Zimmermann et al, EMBO J. 7:367-372 (1988)). The N-terminal extracellular portion of both the Drosophila APPL protein and APP can be secreted via cleavage at or near die membrane (Weidman et al, Cell 57:115-126 (1989); Zimmermann et al, EMBO J. 7:367-372 (1988); and Palmert et al , Proc. Natl. Acad. Sci. USA 56:6338-6342 (1989)). Although d e normal function of the proteins in the APP family remains cryptic, the present results suggest that APLPl , like APP and die Drosophila APPL may be processed in, or reside in, die Golgi. The existence of a family of APP-like proteins implies that these proteins may share a function. The conservation of cysteines at the N-termini is indicative of conserved tertiary and/or quaternary structure, and suggests that these molecules may interact wim a common extracellular molecule. Likewise the strong amino acid conservation within the intracellular C-termini suggests diat die proteins in diis family may interact with a common molecule inside of me cell. A distinct physiological role for APP has yet to be determined. Clues to die function of any of the members of the APP-like family of proteins should help to elucidate die normal function and processing and regulation of APP. Example 3 Mapping the Human Chromosomal Locus Encoding APLPl Portions of the mouse brain cDNA and a 1.8 kb partial cDNA isolated from a human brain cDNA library were used to map the human chromosomal locus encoding APLPl. To determine the best restriction digest for the selective identification of die human chromosomal APLPl fragments, human, mouse and hamster genomic DNAs were analyzed by Southern blot hybridization using die partial human cDNA clone following digestion wid EcoRI, Hindlll, PstI, and Taql. EcoRI was chosen for further analysis since it produced human DNA fragments (approximately 8 kb and 3.3 kb, data not shown) that were clearly discernable from the rodent. A panel of DNAs from 31 human-rodent somatic cell lines (Geissler et al , Somat. Cell Mol. Genet. 77:197-214 (1991)) of known karyotype was digested with EcoRI. These DNAs were then probed with die human APLPl cDNA clone and me hybridization pattern was consistent with the assignment of the APLPl locus to chromosome 19. To determine die regional position of APLPl locus on chromosome 19, the full-lengtii mouse brain APLPl cDNA was hybridized to EcoRI-digested genomic DNA from a number of somatic cell hybrids containing only human chromosome 19 or specific fragments of this autosome as well as otiier chromosomes (G35CCB, G35F3B, GM89A99c7B, G24B2AM, FON1A4, TVBID, 1016A and 5HL94; FIGURE 16). All of these hybrid lines, with the exception of GM89A99c7B, contain the two human specific APLPl bands. GM89A99c7B contains the reciprocal part of the X: 19 translocation occurring in 908K1, G35F3 and G35FCC (FIGURE 16). These results exclude the APLPl locus from the short arm of chromosome 19 and place it between 19ql3.2 and die centromere. Discussion While no physiological role has been determined for APLPl, its map location is interesting in view of its potential relationship to Alzheimer's disease (AD). The chief component of Alzheimer-associated amyloid is die 39-43 amino acid βA4 peptide which is derived from the larger amyloid precursor protein (APP) encoded by a gene on chromosome 21 (Kang et al , Nature 325:733-736 (1987); Robakis et al , Proc. Natl. Acad. Sci. USA 54:4190-4194 (1987); Tanzi et al , Science 235:880-884 (1987)). The gene defect for an early-onset (>65 years of age) form of familial Alzheimer's disease (FAD) has been mapped to chromosome 21 (St. George-Hyslop et al , Science 235:885- 889 (1987)) and a small percentage (<3%) of FAD appears to be caused by mutations within the APP gene (Chartier-Harlin et al , Nature 353:884-846 (1991); Goate et al. , Nature 349:704-706 (1991); Murrell et al. , Science 254:97-99 (1991)). Genetic heterogeneity has also been reported for FAD (St. George-Hyslop et al. , Nature 347: 194-197 (1990)) and a set of late-onset ( > 65 years of age) FAD pedigrees have recently demonstrated linkage to chromosome 19 (Pericak- Vance et al , Am. J. Hum. Genet. 45: 1034-1050 (1991)). Because of the regional chromosomal localization of APLPl to the proximal portion of 19q and die significant homology of diis gene to APP, APLP1 is a candidate for the gene defect responsible for a late-onset form of FAD. Example 4 Isolation and characterization of the Human APLP2 Gene encoding a homologue of the Alzheimer 's Associated Amyloid B protein precursor In an attempt to isolate otiier members of the APP protein family, the mouse APLPl sequence was first used to scan die Genbank database for homologous sequences. In addition to obtaining matches for APP, APPL, and the partial cDNA from rate testes, a match widi an anonymous 274 base pair human brain cDNA entry (Genbank accession number M78104), was noted. This match, which was significant but not identical to mouse APLPl (63% identity), indicated diat M78104 was a small piece of a cDNA encoding a second APLP. In order to characterize the APP-like gene family in more detail, full lengtii cDNAs for diis second APLP, APLP2, were isolated. The isolation and characterization of APLP2 cDNA clones from human brain, provide further support for die hypotiiesis that APP is a member of a highly conserved gene family. A human brain frontal cortex Lambda Zap II cDNA library (Stratagene) was screened wim a probe consisting of a PCR product generated widi primers designed to amplify a portion of the 274 base pair partial cDNA sequence identified in Genbank. To prepare the probe, a primer set (5'GCAACCGAATGGACAGGGTA 3' (SEQ ID NO: 11) and 5'CAAGGCAGCCAGGTAGTTCTC 3' (SEQ ID NO: 12); see FIGURE 17) was used to amplify a 232 base pair product from a human occipital cortex cDNA library. The PCR product was sequenced to confirm its identity and an internal primer set (5OTAAAGAAGGAATGGGAAGAGGC3' (SEQ ID NO: 13) and 5'CCATCCGACGGCGGTCATTCAGC3' (SEQ ID NO: 14); see HGURE 17) was designed and used to amplify a 185 base pair PCR fragment (SG190) that was used for die human brain library screen. Screening, purification and sequencing of die SG190-positive clones, including a full length cDNA were carried out according to standard conditions (Wasco et al. , Proc. Natl. Acad. Sci. USA 59:10758-10762 (1992)). Human APLP2 is encoded by a 706 amino acid sequence diat is similar to APP and APLPl in overall structure as well as amino acid sequence. APLP2 is 52% identical, 69% similar to APP695 (FIGURE 17) (SEQ ID NOS:29-30) and 43% identical, 63% similar to APLPl . Virtually all of the identified domains and motifs that characterize APP, APPL and APLPl are present in APLP2. Specifically, an N-terminal cysteine-rich region (consisting of 12 cysteines), a novel zinc-binding motif (Bush et al. , Neurobiol Aging 13 (supplement 1):A.331 (1992)), an acidic-rich domain, N-glycosylation sites, a hydrophobic membrane spanning domain and a cytoplasmic domain containing a clathrin binding motif and potential serine/threonine, casine kinase I, II and tyrosine phosphorylation sites are conserved in APLP2 (FIGURE 17). FIGURE 18 (SEQ ID NOS:31-33) shows the amino acids that are identical or conservatively substituted in APLP2, APP and APLPl demonstrating die extremely high degree of conservation among these proteins. Some of these stretches of homologous amino acids shown in FIGURE 18 (SEQ ID NOS: 31- 33) may contain potential consensus motifs that are germane to the function of this protein family. Chromosomal Location of APLP2 Gene To determine the chromosomal location of the APLP2 gene, a cDNA probe was hybridized to a filter containing Hind m-digested DNA from a panel of 43 human-rodent somatic cell hybrid lines containing either individual or specific sets of human chromosomes (Pelletier et al, Genomics 70:1079-1082 (1991); Geissler et al , Som Cell Gen 77:207-214 (1991)). The probe detected specific human APLP2 bands in somatic cell hybrid lines consistent widi die assignment of die APLP2 gene locus to human chromosome 11. Specifically, a positive signal for human APLP2 was obtained in a somatic cell hybrid line containing DNA from chromosome 11 as its only human material, and in a hybrid containing chromosome 11 and tiiree other human chromosomes. During the sequencing of die human APLP2 cDNA clones, a single alternatively spliced form containing an exon encoding a 12 amino acid stretch was identified indicating diat like APP, APLP2 is alternatively transcribed (FIGURE 17). Although a portion of die APLP2 cDNA isolated from mouse embryo contained a KPI domain similar to that in APP, a form of adult human APLP2 that contains such a domain has not yet been detected. Northern Blot Analysis Northern blot analysis of fetal peripheral tissue and adult brain regions demonstrated mat the APLP2 message is approximately 4 kb in size (FIGURE 19). Lighter bands at approximately 3 kb and 2 kb may represent cross-hybridizing messages from other members of the .APP/APLP family, or as of yet unisolated APLP2 alternative transcripts. The APLP2 transcript was detected at varying levels in all peripheral and central nervous system tissues tested and displayed a level and pattern of expression diat is extremely similar to that of APP (FIGURE 19A). Both transcripts are expressed in relatively abundant amounts in brain, heart, and kidney and at lower levels in liver and thymus. However, in contrast to APP, APLP2 is expressed at relatively high levels in the small intestine and lung. To determine e distribution of APLP2 transcript in die adult human brain Northern blot analysis was carried out on mRNA from 11 different brain regions (FIGURE 19B). This same blot had been previously hybridized to APP thus allowing a direct comparison of expression of the two genes (Tanzi et al , Science 235:880-884 (1987); Tanzi et al , Nature 337:528-530 (1988)). The 6/15265 -59- levels of APLP2 mRNA were highest in the temporal association cortex (A20, Tanzi et al, Nature 337:528-530 (1988)), the posterior perisylvian cortex- supramarginal gyri (A40), the anterior perisylvian cortex-opercular gyri (A44) and frontal pole of the cortex (A10). These regions which are particularly affected in die brains of AD patients, normally contain a relatively large amount of APP RNA. Moderate hybridization was detected in the cerebellar cortex and die caudate-putamen. Relatively weaker hybridization was seen in the striate, extrastriate, and motor cortices (A17, A18 and A4), die hippocampus, and die tiialamus. Overall, APLP2 reveals a pattern of expression diat is very similar to that of APP (FIGURE 19B; Tanzi et al, Science 235:880-884 (1987); Tanzi et al , Nature 337:528-530 (1988)), although, some differences were noted. For example, APLP2 is expressed at relatively higher levels tiian APP in thalamus, while APP expression is greater than mat of APLP2 in Brodman area A40. FIGURE 20 shows the result of Northern blot hybridization of APLP2 and APP cDNA probes to RNA derived from normal and fetal brains with Down syndrome (DS), and from normal and adult brains with AD. Altiiough APP expression is higher in the DS samples, APLP2 expression is not significantly changed. This result is not unexpected given die extra copy of chromosome 21 present in DS patients. APP expression is slightly lower in AD versus normal adult cerebellum, and is dramatically decreased in AD frontal cortex relative to normal (FIGURE 20). This decrease in APP expression is probably a reflection of AD-related neuronal loss in these areas which is particularly enhanced in die frontal cortex. Surprisingly, it has presently been found diat although APP expression is somewhat decreased in AD cerebellum compound to normal, APLP2 expression is clearly increased in this same AD cerebellum sample (FIGURE 20). One possibility is that diis may reflect a compensatory increase in APLP2 expression in response to lower levels of APP. It is equally conceivable that increased expression of APLP2 preceded die decrease in APP message. The present inventors have discovered diat APLPs may compete widi APP for factors involved wim maturation and processing (Wasco et al. , Proc. Natl Acad. Sci. USA 59:10758-10762 (1992)). This would require diat the two proteins are produced and processed witiiin the same cell populations. To address this issue, non isotopic in situ hybridization studies designed to localize APLP2 mRNA transcripts within the hippocampal formation, a region that is severely affected in AD, have been employed. It was found diat die mRNA for APLP2 is contained in botii die cell soma and, to some extent, neuronal processes of pyramidal neurons in die hippocampal formation (FIGURE 21). Much less hybridization was observed in smaller interneurons, glial cells, and endothelial cells. The subcellular localization is similar to that seen for APP and APLPl messages using me same in situ hybridization procedure (Tanzi et al , Mol. Brain Res. : in press; Hyman et al , Mol. Brain Res. : in press; Wasco et al, Alzheimer's disease and related disorders 7 92 selected communications (in press)). Moreover, the cellular specificity and regional distribution of die APLP messages are also extremely similar to those of APP indicating mat the APP and APLPs are located within die same sets of neurons in the hippocampal formation. Based on die overall conservation of amino acid sequence and domain structure within die APP gene family, these proteins may share common functions and, perhaps be processed similarly (Wasco et al. , Proc. Natl. Acad. Sci. USA 59:10758-10762 (1992)). Recent data further indicate that APLP2 and APP undergo similar processing (unpublished data). Antibodies to APP, APLPl and APPL recognize proteins in the Golgi (Wasco et al , Proc. Natl. Acad. Sci. USA 59:10758-10762 (1992); Zimmermann et al , EMBO J. 7:367-372 (1988); Palacios et al , Mol. Brain Res. 75:195-206 (1992); Luo et al , J. Neurosci. 70:3849-3861 (1990)). Likewise, APLP2 appears to be associated with d e Golgi apparatus unpublished data. This suggests diat maturation of these proteins in the Golgi very likely involves interaction widi common factors. The apparent similarities in the processing and maturation of APP and APLP2 raises the possibility that altered expression of APLP2 or other APLPs could affect the post-translational modification and metabolism of APP in cells where these genes are co-expressed. If APLP2 or other APLPs were to interfere with die proper maturation (e.g. N- or O-glycosylation) of APP, APP could be rerouted into alternative pathways including diose predisposed to amyloid formation. Along these same lines, if the metabolic machinery responsible for processing APP were overburdened widi members of the APLP family, altered metabolism of APP may occur, perhaps resulting in increased production of amyloidogenic fragments. Therefore, although the APLP2 and APLPl do not contain an Aβ domain, they may still ultimately affect the maturation and/or metabolism of APP. It will be appreciated to tiiose skilled in the art diat die invention can be performed witiiin a wide range of equivalent parameters of composition, concentrations, modes of administration, and conditions without departing from the spirit or scope of the invention or any embodiment thereof. The disclosures of all references, patent applications and patents recited herein are hereby incorporated by reference. SEQUENCE LISTING (1) GENERAL INFORMATION: (i) APPLICANT: The General Hospital Corporation (ii) TITLE OF INVENTION: Methods For Modulating Transcription From The Amyloid β-Protein Precursor (APP) Promoter (iii) NUMBER OF SEQUENCES: 33 (iv) CORRESPONDENCE ADDRESS: (A) ADDRESSEE: STERNE, KESSLER, GOLDSTEIN & FOX P.L.L.C. (B) STREET: 1100 New York Ave. , NW (C) CITY: Washington (D) STATE: DC (E) COUNTRY: USA (F) ZIP: 20005 (v) COMPUTER READABLE FORM: (A) MEDIUM TYPE: Floppy disk (B) COMPUTER: IBM PC compatible (C) OPERATING SYSTEM: PC-DOS/MS-DOS (D) SOFTWARE: Patentln Release #1.0, Version #1.25 (vi) PRIORITY APPLICATION DATA: (A) APPLICATION NUMBER: US 08/339,152 (B) FILING DATE: 10-NOV-1994 (C) CLASSIFICATION: (vii) CURRENT APPLICATION DATA: (A) APPLICATION NUMBER: (To Be Advised) (B) FILING DATE: 09-NOV-1995 (C) CLASSIFICATION: (viii) ATTORNEY/AGENT INFORMATION: (A) NAME: Steffe, Eric K. (B) REGISTRATION NUMBER: 36,688 (C) REFERENCE/DOCKET NUMBER: 0609.412PC00 (ix) TELECOMMUNICATION INFORMATION: (A) TELEPHONE: 202-371-2600 (B) TELEFAX: 202-371-2540 (C) TELEX: 248636 SSK (2) INFORMATION FOR SEQ ID NO:l: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 9 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:l: Gin Gin Leu Arg Glu Leu Gin Arg His 1 5 (2) INFORMATION FOR SEQ ID NO:2: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 29 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2: GGGCCGGATC AGCTGACTCG CCTGGCTCT 29 (2) INFORMATION FOR SEQ ID NO:3: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 22 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3: GCTCAGATCT CGAGCTCGGT AC 22 (2) INFORMATION FOR SEQ ID NO:4: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 29 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:4: GCTTACTGTC AGCTGACTCG CCTGGCTCT 29 (2) INFORMATION FOR SEQ ID NO:5: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 29 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:5: GGGCCGGAAT CGTGCTGTCG CCTGGCTCT 29 (2) INFORMATION FOR SEQ ID NO:6: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 29 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:6: GGGCCGGATC AGCTGACGAT ACCTGTCCG 29 (2) INFORMATION FOR SEQ ID NO:7: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 22 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:7: GATCCAGCTG ACTCATCACT AG 22 (2) INFORMATION FOR SEQ ID NO:8: (i) SEQUENCE CHVRACTERISTICS: (A) LENGTH: 28 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:8: GATCACCAGC TGTGGAATGT GTGTGATC 28 (2) INFORMATION FOR SEQ ID NO:9: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 29 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:9: GGGCCGGATC ACGTGACTCG CCTGGCTCT 29 (2) INFORMATION FOR SEQ ID NO:10: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 19 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:10: GGAAATATAG AAGAAGGAG 19 (2) INFORMATION FOR SEQ ID NO:11: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 20 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:11: GCAACCGAAT GGACAGGGTA 20 (2) INFORMATION FOR SEQ ID NO:12: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 21 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:12: CAAGGCAGCC AGGTAGTTCT C 21 (2) INFORMATION FOR SEQ ID NO:13: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 23 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:13: GTAAAGAAGG AATGGGAAGA GGC 23 (2) INFORMATION FOR SEQ ID NO:14: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 23 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:14: CCATCCGACG GCGGTCATTC AGC 23 (2) INFORMATION FOR SEQ ID NO:15: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 107 base pairs (B) TYPE: nucleic acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear (ii) MOLECULE TYPE: DNA (ix) FEATURE: (A) NAME/KEY: CDS (B) LOCATION: 88..2046 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:15: CGGCACGAGG TGGCGCTGGG AGCTCCTGTC ACCGCTGGGG CCGGGTAGGG GCGGGCGGGA 60 GCGCAGGGAC GTGAGGGCCG AGCGGAC ATG GGG CCC ACC AGC CCC GCC GCT 111 Met Gly Pro Thr Ser Pro Ala Ala 1 5 CGC GGT CAG GGT CGC CGC TGG CGA CCG CCG CTG CCG CTG TTG CTG CCA 159 Arg Gly Gin Gly Arg Arg Trp Arg Pro Pro Leu Pro Leu Leu Leu Pro 10 15 20 CTG TCA TTG CTG CTT CTG CGC GCG CAG CTC GCC GTC GGG AAC CTG GCT 207 Leu Ser Leu Leu Leu Leu Arg Ala Gin Leu Ala Val Gly Asn Leu Ala 25 30 35 40 GTT GGG AGC CCC AGC GCG GCC GAG GCT CCG GGG TCG GCT CAA GTG GCT 255 Val Gly Ser Pro Ser Ala Ala Glu Ala Pro Gly Ser Ala Gin Val Ala 45 50 55 GGA CTA TGT GGG CGT CTA ACC CTT CAC CGG GAC TTG CGC ACC GGC CGC 303 Gly Leu Cys Gly Arg Leu Thr Leu His Arg Asp Leu Arg Thr Gly Arg 60 65 70 TGG GAA CCA GAC CCA CAG CGA TCA CGA CGC TGT CTT CTG GAC CCG CAG 351 Trp Glu Pro Asp Pro Gin Arg Ser Arg Arg Cys Leu Leu Asp Pro Gin 75 80 85 CGC GTG CTG GAG TAC TGC AGA CAG ATG TAC CCC GAG CTG CAC ATA GCA 399 Arg Val Leu Glu Tyr Cys Arg Gin Met Tyr Pro Glu Leu His Ile Ala 90 95 100 CGC GTG GAG CAG GCT GCA CAG GCC ATC CCG ATG GAG CGC TGG TGT GGG 447 Arg Val Glu Gin Ala Ala Gin Ala Ile Pro Met Glu Arg Trp Cys Gly 105 110 115 120 GGT ACC CGG AGT GGC AGA TGC GCC CAC CCC CAC CAT GAG GTT GTG CCC 495 Gly Thr Arg Ser Gly Arg Cys Ala His Pro His His Glu Val Val Pro 125 130 135 TTC CAT TGC CTG CCT GGC GAA TTC GTG AGT GAA GCC CTG CTA GTG CCC 543 Phe His Cys Leu Pro Gly Glu Phe Val Ser Glu Ala Leu Leu Val Pro 140 145 150 GAA GGC TGT CGG TTC TTG CAC CAG GAG CGT ATG GAC CAG TGT GAG AGT 591 Glu Gly Cys Arg Phe Leu His Gin Glu Arg Met Asp Gin Cys Glu Ser 155 160 165 TCA ACC AGG AGG CAT CAG GAG GCT CAG GAG GCC TGC AGC TCT CAG GGC 639 Ser Thr Arg Arg His Gin Glu Ala Gin Glu Ala Cys Ser Ser Gin Gly 170 175 180 CTC ATC CTG CAC GGC TCT GGC ATG CTT TTG CCC TGT GGC TCT GAT CGG 687 Leu Ile Leu His Gly Ser Gly Met Leu Leu Pro Cys Gly Ser Asp Arg 185 190 195 200 TTC CGA GGT GTG GAG TAT GTA TGC TGT CCA CCT CCC GCA ACT CCC AAC 735 Phe Arg Gly Val Glu Tyr Val Cys Cys Pro Pro Pro Ala Thr Pro Asn 205 210 215 CCA TCT GGG ATG GCA GCT GGT GAC CCC TCT ACC CGG TCC TGG CCC CTG 783 Pro Ser Gly Met Ala Ala Gly Asp Pro Ser Thr Arg Ser Trp Pro Leu 220 225 230 GGG GGC AGA GCA GAG GGA GGT GAG GAT GAA GAG GAG GTG GAA TCT TTC 831 Gly Gly Arg Ala Glu Gly Gly Glu Asp Glu Glu Glu Val Glu Ser Phe 235 240 245 CCT CAG CCA GTA GAC GAT TAC TTC GTA GAG CCC CCT CAG GCT GAA GAA 879 Pro Gin Pro Val Asp Asp Tyr Phe Val Glu Pro Pro Gin Ala Glu Glu 250 255 260 GAA GAG GAA GAG GAG GAA GAA AGG GCC CCA CCT CCC AGC TCC CAC ACC 927 Glu Glu Glu Glu Glu Glu Glu Arg Ala Pro Pro Pro Ser Ser His Thr 265 270 275 280 CCT GTC ATG GTT AGC AGA GTC ACT CCC ACC CCA AGG CCT ACT GAT GGT 975 Pro Val Met Val Ser Arg Val Thr Pro Thr Pro Arg Pro Thr Asp Gly 285 290 295 GTG GAT GTT TAC TTT GGC ATG CCT GGG GAA ATC GGC GAG CAT GAG GGT 1023 Val Asp Val Tyr Phe Gly Met Pro Gly Glu Ile Gly Glu His Glu Gly 300 305 310 TTC CTG AGG GCC AAG ATG GAC CTG GAG GAG CGT AGG ATG CGC CAG ATT 1071 Phe Leu Arg Ala Lys Met Asp Leu Glu Glu Arg Arg Met Arg Gin Ile 315 320 325 AAT GAG GTG ATG CGT GAA TGG GCC ATG GCT GAC AGC CAA TCT AAG AAC 1119 Asn Glu Val Met Arg Glu Trp Ala Met Ala Asp Ser Gin Ser Lys Asn 330 335 340 CTG CCA AAG GCG GAC AGG CAG GCC CTG AAT GAG CAC TTC CAG TCC ATT 1167 Leu Pro Lys Ala Asp Arg Gin Ala Leu Asn Glu His Phe Gin Ser Ile 345 350 355 360 CTG CAG ACC CTG GAA GAA CAA GTG TCT GGT GAA CGG CAA CGC CTG GTG 1215 Leu Gin Thr Leu Glu Glu Gin Val Ser Gly Glu Arg Gin Arg Leu Val 365 370 375 GAG ACC CAC GCC ACC AGA GTC ATC GCT CTG ATC AAC GAC CAG CGC CGA 1263 Glu Thr His Ala Thr Arg Val Ile Ala Leu Ile Asn Asp Gin Arg Arg 380 385 390 GCA GCC CTG GAA GGT TTC CTG GCA GCC TTA CAG GGC GAT CCG CCT CAG 1311 Ala Ala Leu Glu Gly Phe Leu Ala Ala Leu Gin Gly Asp Pro Pro Gin 395 400 405 GCT GAG CGA GTT CTG ATG GCC CTG AGG CGC TAC CTG CGC GCC GAG CAG 1359 Ala Glu Arg Val Leu Met Ala Leu Arg Arg Tyr Leu Arg Ala Glu Gin 410 415 420 AAA GAG CAG AGG CAC ACT CTG AGG CAC TAC CAG CAC GTG GCC GCA GTG 1407 Lys Glu Gin Arg His Thr Leu Arg His Tyr Gin His Val Ala Ala Val 425 430 435 440 GAT CCT GAG AAG GCC CAG CAG ATG CGC TTT CAG GTC CAG ACC CAC CTT 1455 Asp Pro Glu Lys Ala Gin Gin Met Arg Phe Gin Val Gin Thr His Leu 445 450 455 CAG GTG ATC GAA GAG CGA ATG AAT CAG AGC CTG GGG CTG CTC GAC CAG 1503 Gin Val Ile Glu Glu Arg Met Asn Gin Ser Leu Gly Leu Leu Asp Gin 460 465 470 AAC CCT CAC CTG GCT CAG GAG CTG CGG CCA CAG ATC CAG GAG CTT CTC 1551 Asn Pro His Leu Ala Gin Glu Leu Arg Pro Gin Ile Gin Glu Leu Leu 475 480 485 CTT GCT GAA CAC TTG GGT CCC AGT GAA CTG GAC GCC TCT GTG CCC GGG 1599 Leu Ala Glu His Leu Gly Pro Ser Glu Leu Asp Ala Ser Val Pro Gly 490 495 500 AGC AGC AGT GAG GAC AAA GGT AGC CTC CAG CCT CCC GAA TCC AAG GAC 1647 Ser Ser Ser Glu Asp Lys Gly Ser Leu Gin Pro Pro Glu Ser Lys Asp 505 510 515 520 GAT CCC CCA GTG ACC CTT CCA AAA GGG TCC ACA GAT CAA GAG TCA TCC 1695 Asp Pro Pro Val Thr Leu Pro Lys Gly Ser Thr Asp Gin Glu Ser Ser 525 530 535 TCC TCT GGG AGA GAG AAG CTA ACT CCA CTG GAG CAG TAT GAG CAA AAG 1743 Ser Ser Gly Arg Glu Lys Leu Thr Pro Leu Glu Gin Tyr Glu Gin Lys 540 545 550 GTG AAT GCA TCC GCC CCG AGG GGG TTT CCG TTC CAC TCG TCA GAT ATC 1791 Val Asn Ala Ser Ala Pro Arg Gly Phe Pro Phe His Ser Ser Asp Ile 555 560 565 CAG CGG GAT GAA CTG GCT CCT TCC GGG ACT GGA GTG TCC CGA GAG GCC 1839 Gin Arg Asp Glu Leu Ala Pro Ser Gly Thr Gly Val Ser Arg Glu Ala 570 575 580 TTG TCA GGT CTG CTG ATC ATG GGA GCT GGA GGA GGC TCT CTC ATT GTC 1887 Leu Ser Gly Leu Leu Ile Met Gly Ala Gly Gly Gly Ser Leu Ile Val 585 590 595 600 CTA TCC TTG CTG CTT CTG CGC AAG AAG AAA CCC TAT GGG ACT ATC AGC 1935 Leu Ser Leu Leu Leu Leu Arg Lys Lys Lys Pro Tyr Gly Thr Ile Ser 605 610 615 CAT GGA GTG GTG GAG GTG GAC CCC ATG CTG ACC CTG GAG GAG CAG CAG 1983 His Gly Val Val Glu Val Asp Pro Met Leu Thr Leu Glu Glu Gin Gin 620 625 630 CTC CGG GAA CTT CAG AGG CAT GGC TAT GAG AAC CCC ACC TAC CGC TTC 2031 Leu Arg Glu Leu Gin Arg His Gly Tyr Glu Asn Pro Thr Tyr Arg Phe 635 640 645 CTG GAA GAA CGA CCT TGACCCCTAC CCTAGCTGCC TTCAGCTGAG CCCTACTGCC 20B6 Leu Glu Glu Arg Pro 650 TTCTTCCGGC CCCCCAAACC CAACTCCCAG CTTCCGGTGG GGGAGGGAGA TCTTGACAAA 2146 TTCATTCTTG TTTCCCCTTC CTAGTTCCAA ATTCCACACC CTTAGAAATC CCCAGCTCCT 2206 GTCCCACAAG GGACCTCTTC ACCTTAATTT ATTTTACGTT AATTTATTGC TCCTTAAGGT 2266 GACCTGGGTC CCAGGTATGT ATGTCACTCC CTGGAATTCA CCATCCCACG TTTCTTCACT 2326 AACATCCCAA TAAACTCCTC TTTCCCTCCG GC 2358 (2) INFORMATION FOR SEQ ID NO:16: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 653 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) MOLECULE TYPE: protein (xi) SEQUENCE DESCRIPTION: SEQ ID NO:16: Met Gly Pro Thr Ser Pro Ala Ala Arg Gly Gin Gly Arg Arg Trp Arg 1 5 10 15 Pro Pro Leu Pro Leu Leu Leu Pro Leu Ser Leu Leu Leu Leu Arg Ala 20 25 30 Gin Leu Ala Val Gly Asn Leu Ala Val Gly Ser Pro Ser Ala Ala Glu 35 40 45 Ala Pro Gly Ser Ala Gin Val Ala Gly Leu Cys Gly Arg Leu Thr Leu 50 55 60 His Arg Asp Leu Arg Thr Gly Arg Trp Glu Pro Asp Pro Gin Arg Ser 65 70 75 80 Arg Arg Cys Leu Leu Asp Pro Gin Arg Val Leu Glu Tyr Cys Arg Gin 85 90 95 Met Tyr Pro Glu Leu His Ile Ala Arg Val Glu Gin Ala Ala Gin Ala 100 105 110 lie Pro Met Glu Arg Trp Cys Gly Gly Thr Arg Ser Gly Arg Cys Ala 115 120 125 His Pro His His Glu Val Val Pro Phe His Cys Leu Pro Gly Glu Phe 130 135 140 Val Ser Glu Ala Leu Leu Val Pro Glu Gly Cys Arg Phe Leu His Gin 145 150 155 160 Glu Arg Met Asp Gin Cys Glu Ser Ser Thr Arg Arg His Gin Glu Ala 165 170 175 Gin Glu Ala Cys Ser Ser Gin Gly Leu Ile Leu His Gly Ser Gly Met 180 185 190 Leu Leu Pro Cys Gly Ser Asp Arg Phe Arg Gly Val Glu Tyr Val Cys 195 200 205 Cys Pro Pro Pro Ala Thr Pro Asn Pro Ser Gly Met Ala Ala Gly Asp 210 215 220 Pro Ser Thr Arg Ser Trp Pro Leu Gly Gly Arg Ala Glu Gly Gly Glu 225 230 235 240 Asp Glu Glu Glu Val Glu Ser Phe Pro Gin Pro Val Asp Asp Tyr Phe 245 250 255 Val Glu Pro Pro Gin Ala Glu Glu Glu Glu Glu Glu Glu Glu Glu Arg 260 265 270 Ala Pro Pro Pro Ser Ser His Thr Pro Val Met Val Ser Arg Val Thr 275 280 285 Pro Thr Pro Arg Pro Thr Asp Gly Val Asp Val Tyr Phe Gly Met Pro 290 295 300 Gly Glu Ile Gly Glu His Glu Gly Phe Leu Arg Ala Lys Met Asp Leu 305 310 315 320 Glu Glu Arg Arg Met Arg Gin Ile Asn Glu Val Met Arg Glu Trp Ala 325 330 335 Met Ala Asp Ser Gin Ser Lys Asn Leu Pro Lys Ala Asp Arg Gin Ala 340 345 350 Leu Asn Glu His Phe Gin Ser Ile Leu Gin Thr Leu Glu Glu Gin Val 355 360 365 Ser Gly Glu Arg Gin Arg Leu Val Glu Thr His Ala Thr Arg Val Ile 370 375 380 Ala Leu Ile Asn Asp Gin Arg Arg Ala Ala Leu Glu Gly Phe Leu Ala 385 390 395 400 Ala Leu Gin Gly Asp Pro Pro Gin Ala Glu Arg Val Leu Met Ala Leu 405 410 415 Arg Arg Tyr Leu Arg Ala Glu Gin Lys Glu Gin Arg His Thr Leu Arg 420 425 430 His Tyr Gin His Val Ala Ala Val Asp Pro Glu Lys Ala Gin Gin Met 435 440 445 Arg Phe Gin Val Gin Thr His Leu Gin Val Ile Glu Glu Arg Met Asn 450 455 460 Gin Ser Leu Gly Leu Leu Asp Gin Asn Pro His Leu Ala Gin Glu Leu 465 470 475 480 Arg Pro Gin Ile Gin Glu Leu Leu Leu Ala Glu His Leu Gly Pro Ser 485 490 495 Glu Leu Asp Ala Ser Val Pro Gly Ser Ser Ser Glu Asp Lys Gly Ser 500 505 510 Leu Gin Pro Pro Glu Ser Lys Asp Asp Pro Pro Val Thr Leu Pro Lys 515 520 525 Gly Ser Thr Asp Gin Glu Ser Ser Ser Ser Gly Arg Glu Lys Leu Thr 530 535 540 Pro Leu Glu Gin Tyr Glu Gin Lys Val Asn Ala Ser Ala Pro Arg Gly 545 550 555 560 Phe Pro Phe His Ser Ser Asp Ile Gin Arg Asp Glu Leu Ala Pro Ser 565 570 575 Gly Thr Gly Val Ser Arg Glu Ala Leu Ser Gly Leu Leu Ile Met Gly 580 585 590 Ala Gly Gly Gly Ser Leu Ile Val Leu Ser Leu Leu Leu Leu Arg Lys 595 600 605 Lys Lys Pro Tyr Gly Thr Ile Ser His Gly Val Val Glu Val Asp Pro 610 615 620 Met Leu Thr Leu Glu Glu Gin Gin Leu Arg Glu Leu Gin Arg His Gly 625 630 635 640 Tyr Glu Asn Pro Thr Tyr Arg Phe Leu Glu Glu Arg Pro 645 650 (2) INFORMATION FOR SEQ ID NO:17: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 634 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:17: Leu Leu Leu Pro Leu Ser Leu Leu Leu Leu Arg Ala Gin Leu Ala Val 1 5 10 15 Gly Asn Leu Ala Val Gly Ser Pro Ser Ala Ala Glu Ala Pro Gly Ser 20 25 30 Ala Gin Val Ala Gly Leu Cys Gly Arg Leu Thr Leu His Arg Asp Leu 35 40 45 Arg Thr Gly Arg Trp Glu Pro Asp Pro Gin Arg Ser Arg Arg Cys Leu 50 55 60 Leu Asp Pro Gin Arg Val Leu Glu Tyr Cys Arg Gin Met Tyr Pro Glu 65 70 75 80 Leu His Ile Ala Arg Val Glu Gin Ala Ala Gin Ala Ile Pro Met Glu 85 90 95 Arg Trp Cys Gly Gly Thr Arg Ser Gly Arg Cys Ala His Pro His His 100 105 110 Glu Val Val Pro Phe His Cys Leu Pro Gly Glu Phe Val Ser Glu Ala 115 120 125 Leu Leu Val Pro Glu Gly Cys Arg Phe Leu His Gin Glu Arg Met Asp 130 135 140 Gin Cys Glu Ser Ser Thr Arg Arg His Gin Glu Ala Gin Glu Ala Cys 145 150 155 160 Ser Ser Gin Gly Leu Ile Leu His Gly Ser Gly Met Leu Leu Pro Cys 165 170 175 Gly Ser Asp Arg Phe Arg Gly Val Glu Tyr Val Cys Cys Pro Pro Pro 180 185 190 Ala Thr Pro Asn Pro Ser Gly Met Ala Ala Gly Asp Pro Ser Thr Arg 195 200 205 Ser Trp Pro Leu Gly Gly Arg Ala Glu Gly Gly Glu Asp Glu Glu Glu 210 215 220 Val Glu Ser Phe Pro Gin Pro Val Asp Asp Tyr Phe Val Glu Pro Pro 225 230 235 240 Gin Ala Glu Glu Glu Glu Glu Glu Glu Glu Glu Arg Ala Pro Pro Pro 245 250 255 Ser Ser His Thr Pro Val Met Val Ser Arg Val Thr Pro Thr Pro Arg 260 265 270 Pro Thr Asp Gly Val Asp Val Tyr Phe Gly Met Pro Gly Glu Ile Gly 275 280 285 Glu His Glu Gly Phe Leu Arg Ala Lys Met Asp Leu Glu Glu Arg Arg 290 295 300 Met Arg Gin Ile Asn Glu Val Met Arg Glu Trp Ala Met Ala Asp Ser 305 310 315 320 Gin Ser Lys Asn Leu Pro Lys Ala Asp Arg Gin Ala Leu Asn Glu His 325 330 335 Phe Gin Ser Ile Leu Gin Thr Leu Glu Glu Gin Val Ser Gly Glu Arg 340 345 350 Gin Arg Leu Val Glu Thr His Ala Thr Arg Val Ile Ala Leu Ile Asn 355 360 365 Asp Gin Arg Arg Ala Ala Leu Glu Gly Phe Leu Ala Ala Leu Gin Gly 370 375 380 Asp Pro Pro Gin Ala Glu Arg Val Leu Met Ala Leu Arg Arg Tyr Leu 385 390 395 400 Arg Ala Glu Gin Lys Glu Gin Arg His Thr Leu Arg His Tyr Gin His 405 410 415 Val Ala Ala Val Asp Pro Glu Lys Ala Gin Gin Met Arg Phe Gin Val 420 425 430 Gin Thr His Leu Gin Val Ile Glu Glu Arg Met Asn Gin Ser Leu Gly 435 440 445 Leu Leu Asp Gin Asn Pro His Leu Ala Gin Glu Leu Arg Pro Gin Ile 450 455 460 Gin Glu Leu Leu Leu Ala Glu His Leu Gly Pro Ser Glu Leu Asp Ala 465 470 475 480 Ser Val Pro Gly Ser Ser Ser Glu Asp Lys Gly Ser Leu Gin Pro Pro 485 490 495 Glu Ser Lys Asp Asp Pro Pro Val Thr Leu Pro Lys Gly Ser Thr Asp 500 505 510 Gin Glu Ser Ser Ser Ser Gly Arg Glu Lys Leu Thr Pro Leu Glu Gin 515 520 525 Tyr Glu Gin Lys Val Asn Ala Ser Ala Pro Arg Gly Phe Pro Phe His 530 535 540 Ser Ser Asp Ile Gin Arg Asp Glu Leu Ala Pro Ser Gly Thr Gly Val 545 550 555 560 Ser Arg Glu Ala Leu Ser Gly Leu Leu Ile Met Gly Ala Gly Gly Gly 565 570 575 Ser Leu Ile Val Leu Ser Leu Leu Leu Leu Arg Lys Lys Lys Pro Tyr 580 585 590 Gly Thr Ile Ser His Gly Val Val Glu Val Asp Pro Met Leu Thr Leu 595 600 605 Glu Glu Gin Gin Leu Arg Glu Leu Gin Arg His Gly Tyr Glu Asn Pro 610 615 620 Thr Tyr Arg Phe Leu Glu Glu Glu Arg Pro 625 630 (2) INFORMATION FOR SEQ ID NO:18: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 694 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:18: Met Leu Pro Gly Leu Ala Leu Leu Leu Leu Ala Ala Trp Thr Ala Arg 1 5 10 15 Ala Leu Glu Val Pro Thr Asp Gly Asn Ala Gly Leu Leu Ala Glu Pro 20 25 30 Gin Ile Ala Met Phe Cys Gly Arg Leu Asn Met His Met Asn Val Gin 35 40 45 Asn Gly Lys Trp Asp Ser Asp Pro Ser Gly Thr Lys Thr Cys Ile Asp 50 55 60 Thr Lys Glu Gly Ile Leu Gin Tyr Cys Gin Glu Val Tyr Pro Glu Leu 65 70 75 80 Gin Ile Thr Asn Val Val Glu Ala Asn Gin Pro Val Thr Ile Gin Asn 85 90 95 Trp Cys Lys Arg Gly Arg Lys Gin Cys Lys Thr His Pro His Phe Val 100 105 HO Ile Pro Tyr Arg Cys Leu Val Gly Glu Phe Val Ser Asp Ala Leu Leu 115 120 125 Val Pro Asp Lys Cys Lys Phe Leu His Gin Glu Arg Met Asp Val Cys 130 135 140 -76- Glu Thr His Leu His Trp His Thr Val Ala Lys Glu Thr Cys Ser Glu 145 150 155 160 Lys Ser Thr Asn Leu His Asp Tyr Gly Met Leu Leu Pro Cys Gly Ile 165 170 175 Asp Lys Phe Arg Gly Val Glu Phe Val Cys Cys Pro Leu Ala Glu Glu 180 185 190 Ser Asp Asn Val Asp Ser Ala Asp Ala Glu Glu Asp Asp Ser Asp Val 195 200 205 Trp Trp Gly Gly Ala Asp Thr Asp Tyr Ala Asp Gly Ser Glu Asp Lys 210 215 220 Val Val Glu Val Ala Glu Glu Glu Glu Val Ala Glu Val Glu Glu Glu 225 230 235 240 Glu Ala Asp Asp Asp Glu Asp Asp Glu Asp Gly Asp Glu Val Glu Glu 245 250 255 Glu Ala Glu Glu Pro Tyr Glu Glu Ala Thr Glu Arg Thr Thr Ser Ile 260 265 270 Ala Thr Thr Thr Thr Thr Thr Thr Glu Ser Val Glu Glu Val Val Arg 275 280 285 Val Pro Thr Thr Ala Ala Ser Thr Pro Asp Ala Val Asp Lys Tyr Leu 290 295 300 Glu Thr Pro Gly Asp Glu Asn Glu His Ala His Phe Gin Lys Ala Lys 305 310 315 320 Glu Arg Leu Glu Ala Lys His Arg Glu Arg Met Ser Gin Val Met Arg 325 330 335 Glu Trp Glu Glu Ala Glu Arg Gin Ala Lys Asn Leu Pro Lys Ala Asp 340 345 350 Lys Lys Ala Val Ile Gin His Phe Gin Glu Lys Val Glu Ser Leu Glu 355 360 365 Gin Glu Ala Ala Asn Glu Arg Gin Gin Leu Val Glu Thr His Met Ala 370 375 380 Arg Val Glu Ala Met Leu Asn Asp Arg Arg Arg Leu Ala Leu Glu Asn 385 390 395 400 Tyr Ile Thr Ala Leu Gin Ala Val Pro Pro Arg Pro Arg His Val Phe 405 410 415 Asn Met Leu Lys Lys Tyr Val Arg Ala Glu Gin Lys Asp Arg Gin His 420 425 430 Thr Leu Lys His Phe Glu His Val Arg Met Val Asp Pro Lys Lys Ala 435 440 445 Ala Gin Ile Arg Ser Gin Val Met Thr His Leu Arg Val Ile Tyr Glu 450 455 460 Arg Met Asn Gin Ser Leu Ser Leu Leu Tyr Asn Val Pro Ala Val Ala 465 470 475 480 Glu Glu Ile Gin Asp Glu Val Asp Glu Leu Leu Gin Lys Glu Gin Asn 485 490 495 Tyr Ser Asp Asp Val Leu Ala Asn Met Ile Ser Glu Pro Arg Ile Ser 500 505 510 Tyr Gly Asn Asp Ala Leu Met Pro Ser Leu Thr Glu Thr Lys Thr Thr 515 520 525 Val Glu Leu Leu Pro Val Asn Gly Glu Phe Ser Leu Asp Asp Leu Gin 530 535 540 Pro Trp His Ser Phe Gly Ala Asp Ser Val Pro Ala Asn Thr Glu Asn 545 550 555 560 Glu Val Glu Pro Val Asp Ala Arg Pro Ala Ala Asp Arg Gly Leu Thr 565 570 575 Thr Arg Pro Gly Ser Gly Leu Thr Asn Ile Lys Thr Glu Glu Ile Ser 580 585 590 Glu Val Lys Met Asp Ala Glu Phe Arg His Asp Ser Gly Tyr Glu Val 595 600 605 His His Gin Lys Leu Val Phe Phe Ala Glu Asp Val Gly Ser Asn Lys 610 615 620 Gly Ala Ile Ile Gly Leu Met Val Gly Gly Val Val Ile Ala Thr Val 625 630 635 640 Ile Val Ile Thr Leu Val Met Leu Lys Lys Lys Gin Tyr Thr Ser Ile 645 650 655 His His Gly Val Val Glu Val Asp Ala Ala Val Thr Pro Glu Glu Arg 660 665 670 His Leu Ser Lys Met Gin Gin Asn Gly Tyr Glu Asn Pro Thr Tyr Lys 675 680 685 Phe Phe Glu Gin Met Gin 690 (2) INFORMATION FOR SEQ ID NO:19: -78- (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 190 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:19: Leu Leu Leu Pro Leu Ser Leu Leu Leu Leu Arg Ala Gin Leu Ala Val 1 5 10 15 Gly Asn Leu Ala Val Gly Ser Pro Ser Ala Ala Glu Ala Pro Gly Ser 20 25 30 Ala Gin Val Ala Gly Leu Cys Gly Arg Leu Thr Leu His Arg Asp Leu 35 40 45 Arg Thr Gly Arg Trp Glu Pro Asp Pro Gin Arg Ser Arg Arg Cys Leu 50 55 60 Leu Asp Pro Gin Arg Val Leu Glu Tyr Cys Arg Gin Met Tyr Pro Glu 65 70 75 80 Leu His Ile Ala Arg Val Glu Gin Ala Ala Gin Ala Ile Pro Met Glu 85 90 95 Arg Trp Cys Gly Gly Thr Arg Ser Gly Arg Cys Ala His Pro His His 100 105 110 Glu Val Val Pro Phe His Cys Leu Pro Gly Glu Phe Val Ser Glu Ala 115 120 125 Leu Leu Val Pro Glu Gly Cys Arg Phe Leu His Gin Glu Arg Met Asp 130 135 140 Gin Cys Glu Ser Ser Thr Arg Arg His Gin Glu Ala Gin Glu Ala Cys 145 150 155 160 Ser Ser Gin Gly Leu Ile Leu His Gly Ser Gly Met Leu Leu Pro Cys 165 170 175 Gly Ser Asp Arg Phe Arg Gly Val Glu Tyr Val Cys Cys Pro 180 185 190 (2) INFORMATION FOR SEQ ID NO:20: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 188 amino acids (B) TYPE: amino acid (C) STRANDEDNESS: single (D) TOPOLOGY: linear ( i) SEQUENCE DESCRIPTION: SEQ ID NO:20: Met Leu Pro Gly Leu Ala Leu Leu Leu Leu Ala Ala Trp Thr Ala Arg 1 5 10 15 Ala Leu Glu Val Pro Thr Asp Gly Asn Ala Gly Leu Leu Ala Glu Pro 20 25 30 Gin Ile Ala Met Phe Cys Gly Arg Leu Asn Met His Asn Met Val Gin 35 40 45 Asn Gly Lys Trp Asp Ser Asp Pro Ser Gly Thr Lys Thr Cys Ile Asp 50 55 60 Thr Lys Glu Gly Ile Leu Gin Tyr Cys Gin Glu Val Tyr Pro Glu Leu 65 70 75 80 Gin Ile Thr Asn Val Val Glu Ala Asn Gin Pro Val Thr Ile Gin Asn 85 90 95 Trp Cys Lys Arg Gly Arg Lys Gin Cys Lys Thr His Pro His Phe Val 100 105 110 Ile Pro Tyr Arg Cys Leu Val Gly Glu Phe Val Ser Asp Ala Leu Leu 115 120 125 Val Pro Asp Lys Cys Lys Phe Leu His Gin Glu Arg Met Asp Val Cys 130 135 140 Glu Thr His Leu His Trp His Thr Val Ala Lys Glu Thr Cys Ser Glu 145 150 155 160 Lys Ser Thr Asn Leu His Asp Tyr Gly Met Leu Leu Pro Cys Gly Ile 165 170 175 Asp Lys Phe Arg Gly Val Glu Phe Val Cys Cys Pro 180 185 (2) INFORMATION FOR SEQ ID NO:21: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 197 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:21: Met Cys Ala Ala Leu Arg Arg Asn Leu Leu Leu Arg Ser Leu Trp Val 1 5 10 15 Val Leu Ala Ile Gly Thr Ala Gin Val Gin Ala Ala Ser Ser Pro Arg 20 25 30 Trp Pro Gin Ile Ala Val Leu Cys Glu Ala Gly Gin Ile Tyr Gin Pro 35 40 45 Gin Tyr Leu Ser Glu Glu Gly Arg Trp Val Thr Asp Leu Ser Lys Lys 50 55 60 Thr Thr Gly Pro Thr Cys Leu Arg Asp Lys Met Asp Leu Asp Tyr Cys 65 70 75 80 Lys Lys Ala Tyr Pro Asn Arg Asp Ile Thr Asn Ile Val Glu Ser Ser 85 90 95 His Tyr Gin Lys Ile Gly Gly Trp Cys Arg Gin Gly Ala Leu Asn Ala 100 105 110 Ala Lys Cys Lys Gly Ser His Arg Trp Ile Lys Pro Phe Arg Cys Leu 115 120 125 Gly Pro Phe Gin Ser Asp Ala Leu Leu Tyr Pro Glu Gly Cys Leu Phe 130 135 140 Asp His Ile His Asn Ala Ser Arg Cys Trp Pro Phe Val Arg Trp Asn 145 150 155 160 Gin Thr Gly Ala Ala Ala Cys Gin Glu Arg Gly Met Gly Met Arg Thr 165 170 175 Phe Ala Met Leu Leu Pro Cys Gly Ile Ser Val Phe Ser Gly Val Glu 180 185 190 Phe Val Cys Cys Pro 195 (2) INFORMATION FOR SEQ ID NO:22: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 155 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:22: Ala Lys Met Asp Leu Glu Glu Arg Arg Met Arg Gin Ile Asn Glu Val 1 5 10 15 Met Arg Glu Trp Ala Met Ala Asp Ser Gin Ser Lys Asn Leu Pro Lys 20 25 30 Ala Asp Arg Gin Ala Leu Asn Glu His Phe Gin Ser Ile Leu Gin Thr 35 40 45 Leu Glu Glu Gin Val Ser Gly Glu Arg Gin Arg Leu Val Glu Thr His 50 55 60 Ala Thr Arg Val Ile Ala Leu Ile Asn Asp Gin Arg Arg Ala Ala Leu 65 70 75 80 Glu Gly Phe Leu Ala Ala Leu Gin Gly Asp Pro Pro Gin Ala Glu Arg 85 90 95 Val Leu Met Ala Leu Arg Arg Tyr Leu Arg Ala Glu Gin Lys Glu Gin 100 105 110 Arg His Thr Leu Arg His Tyr Gin His Val Ala Ala Val Asp Pro Glu 115 120 125 Lys Ala Gin Gin Met Arg Phe Gin Val Gin Thr His Leu Gin Val Ile 130 135 140 Glu Glu Arg Met Asn Gin Ser Leu Gly Leu Leu 145 150 155 (2) INFORMATION FOR SEQ ID NO:23: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 155 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:23: Ala Lys Glu Arg Leu Glu Ala Lys His Arg Glu Arg Met Ser Gin Val 1 5 10 15 Met Arg Glu Trp Glu Glu Ala Glu Arg Gin Ala Lys Asn Leu Pro Lys 20 25 30 Ala Asp Lys Lys Ala Val Ile Gin His Phe Gin Glu Lys Val Glu Ser 35 40 45 Leu Glu Gin Glu Ala Ala Asn Glu Arg Gin Gin Leu Val Glu Thr His 50 55 60 Met Ala Arg Val Glu Ala Met Leu Asn Asp Arg Arg Arg Leu Ala Leu 65 70 75 80 Glu Asn Tyr Ile Thr Ala Leu Gin Ala Val Pro Pro Arg Pro Arg His 85 90 95 Val Phe Asn Met Leu Lys Lys Tyr Val Arg Ala Glu Gin Lys Asp Arg 100 105 110 Gin His Thr Leu Lys His Phe Glu His Val Arg Met Val Asp Pro Lys 115 120 125 Lys Ala Ala Gin Ile Arg Ser Gin Val Met Thr His Leu Arg Val Ile 130 135 140 Tyr Glu Arg Met Asn Gin Ser Leu Ser Leu Leu 145 150 155 (2) INFORMATION FOR SEQ ID NO:24: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 166 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:24: Ser Gin Lys Arg Leu Glu Glu Ser His Arg Glu Lys Val Thr Arg Val 1 5 10 15 Met Lys Asp Trp Ser Asp Leu Glu Glu Lys Tyr Gin Asp Met Arg Leu 20 25 30 Ala Asp Pro Lys Ala Ala Gin Ser Phe Lys Gin Arg Met Thr Ala Arg 35 40 45 Phe Gin Thr Ser Val Gin Ala Leu Glu Glu Glu Gly Asn Ala Glu Lys 50 55 60 His Gin Leu Ala Ala Met His Gin Gin Arg Val Leu Ala His Ile Asn 65 70 75 80 Gin Arg Lys Arg Glu Ala Met Thr Cys Tyr Thr Gin Ala Leu Thr Glu 85 90 95 Gin Pro Pro Asn Ala His His Val Glu Lys Cys Leu Gin Lys Ile Leu 100 105 110 Arg Ala Leu His Lys Asp Arg Ala His Ala Leu Ala His Tyr Arg His 115 120 125 Leu Leu Asn Ser Gly Gly Pro Gly Gly Leu Glu Ala Ala Ala Ser Glu 130 135 140 Arg Pro Arg Thr Leu Glu Arg Leu Ile Asp Ile Asp Arg Ala Val Asn 145 150 155 160 Gln Ser Met Thr Met Leu 165 (2) INFORMATION FOR SEQ ID NO:25: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 46 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:25: Lys Lys Lys Pro Tyr Gly Thr Ile Ser His Gly Val Val Glu Val Asp 1 5 10 15 Pro Met Leu Thr Leu Glu Glu Gin Gin Leu Arg Glu Leu Gin Arg His 20 25 30 Gly Tyr Glu Asn Pro Thr Tyr Arg Phe Leu Glu Glu Arg Pro 35 40 45 (2) INFORMATION FOR SEQ ID NO:26: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 47 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:26: Lys Lys Lys Gin Tyr Thr Ser Ile His His Gly Val Val Glu Val Asp 1 5 10 15 Ala Ala Val Thr Pro Glu Glu Arg His Leu Ser Lys Met Gin Gin Asn 20 25 30 Gly Tyr Glu Asn Pro Thr Tyr Lys Phe Phe Glu Gin Met Gin Asn 35 40 45 (2) INFORMATION FOR SEQ ID NO:27: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 53 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear ( i) SEQUENCE DESCRIPTION: SEQ ID NO:27: Lys Trp Arg Thr Ser Arg Ser Pro His Ala Gin Gly Phe Ile Glu Val 1 5 10 15 Asp Gin Asn Val Thr Thr His His Pro Ile Val Arg Glu Glu Lys Ile 20 25 30 Val Pro Asn Met Gin Ile Asn Gly Tyr Glu Asn Pro Thr Tyr Lys Tyr 35 40 45 Phe Glu Val Lys Glu 50 (2) INFORMATION FOR SEQ ID NO:28: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 47 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (Xi) SEQUENCE DESCRIPTION: SEQ ID NO:28: Arg Lys Arg Gin Tyr Gly Thr Ile Ser His Gly Ile Val Glu Val Asp 1 5 10 15 Pro Met Leu Thr Pro Glu Glu Arg His Leu Asn Lys Met Gin Asn His 20 25 30 Gly Tyr Glu Asn Pro Thr Tyr Lys Tyr Leu Glu Gin Met Gin Ile 35 40 45 (2) INFORMATION FOR SEQ ID NO:29: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 706 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:29: Met Ala Ala Thr Gly Thr Ala Ala Arg Ala Ala Thr Gly Arg Leu Leu 1 5 10 15 Leu Leu Leu Leu Val Gly Leu Thr Ala Pro Ala Ala Ala Leu Ala Gly 20 25 30 Tyr Ile Glu Ala Leu Ala Ala Ala Ala Gly Thr Gly Phe Ala Val Ala 35 40 45 Glu Pro Gin Ile Ala Met Phe Cys Gly Lys Leu Asn Met His Val Asn 50 55 60 Ile Gin Thr Gly Lys Trp Glu Pro Asp Pro Thr Gly Thr Lys Ser Cys 65 70 75 80 Phe Arg Thr Lys Glu Glu Val Leu Gin Tyr Cys Gin Glu Met Tyr Pro 85 90 95 Glu Leu Gin Ile Thr Asn Val Met Glu Ala Asn Gin Arg Val Ser Ile 100 105 110 Asp Asn Trp Cys Arg Arg Asp Lys Lys Gin Cys Lys Ser Arg Phe Val 115 120 125 Thr Pro Phe Lys Cys Leu Val Gly Glu Phe Val Ser Asp Val Leu Leu 130 135 140 Val Pro Glu Lys Cys Arg Phe Phe His Lys Glu Arg Met Glu Val Cys 145 150 155 160 Glu Asn His Gin His Trp His Thr Val Val Lys Glu Ala Cys Leu Thr 165 170 175 Gin Gly Met Thr Leu Tyr Ser Tyr Gly Met Leu Leu Pro Cys Gly Val 180 185 190 Asp Gin Phe His Gly Thr Glu Tyr Val Cys Cys Pro Gin Thr Lys Asp 195 200 205 Tyr Trp Ser Val Ser Lys Glu Glu Glu Glu Glu Glu Asp Glu Glu Glu 210 215 220 Glu Glu Glu Glu Asp Glu Glu Glu Asp Tyr Asp Val Tyr Lys Ser Glu 225 230 235 240 Phe Pro Thr Glu Ala Asp Leu Glu Asp Phe Thr Glu Ala Ala Val Asp 245 250 255 Glu Asp Asp Glu Asp Glu Glu Glu Gly Glu Glu Val Val Glu Asp Arg 260 265 270 Asp Tyr Tyr Tyr Asp Thr Phe Lys Gly Asp Asp Tyr Asn Glu Glu Asn 275 280 285 Pro Thr Glu Pro Gly Ser Asp Gly Thr Met Ser Asp Lys Glu Ile Thr 290 295 300 His Asp Val Lys Val Pro Pro Thr Pro Leu Pro Thr Asn Asp Val Asp 305 310 315 320 Val Tyr Phe Glu Thr Ser Ala Asp Asp Asn Glu His Ala Arg Phe Gin 325 330 335 Lys Ala Glu Lys Glu Gin Leu Ile Glu Arg His Arg Asn Arg Met Asp 340 345 350 Arg Val Lys Lys Glu Trp Glu Glu Ala Glu Leu Gin Ala Lys Asn Leu 355 360 365 Pro Lys Ala Glu Arg Gin Thr Leu Ile Gin His Phe Gin Ala Met Val 370 375 380 Lys Ala Leu Glu Lys Ala Glu Ala Ala Ser Glu Lys Gin Gin Leu Val 385 390 395 400 Glu Thr His Leu Ala Arg Val Glu Ala Met Leu Asn Asp Arg Arg Met 405 410 415 Ala Leu Glu Asn Tyr Leu Ala Ala Leu Gin Arg Ser Asp Pro Pro Arg 420 425 430 Pro His Arg Ile Leu Gin Pro Leu Arg Arg Tyr Val Arg Ala Glu Asn 435 440 445 Lys Asp Arg Leu His Thr Ile Arg His Tyr Gin His Val Leu Ala Val 450 455 460 Asp Pro Glu Lys Ala Ala Gin Met Lys Ser Gin Val Met Thr His Leu 465 470 475 480 His Val Ile Glu Glu Arg Arg Asn Gin Ser Leu Ser Leu Leu Tyr Lys 485 490 495 Asp Pro Tyr Val Ala Arg Ile Gin Glu Asn Asp Glu Leu Leu Gin Ala 500 505 510 Glu Arg Ala Asp Met Asp Gin Phe Thr Ala Ser Ile Ser Glu Thr Pro 515 520 525 Val Asp Val Arg Val Ser Ser Glu Glu Ser Glu Glu Ile Pro Pro Phe 530 535 540 His Pro Phe His Pro Phe Pro Ala Leu Pro Glu Asn Glu Asp Thr Gin 545 550 555 560 Pro Glu Leu Tyr His Pro Met Lys Lys Gly Ser Gly Val Gly Glu Gin 565 570 575 Asp Gly Gly Leu Ile Gly Ala Glu Glu Lys Val Ile Asn Ser Lys Asn 580 585 590 Lys Val Asp Glu Asn Met Val Ile Asp Glu Thr Leu Asp Lys Glu Met 595 600 605 lie Phe Asn Ala Glu Arg Val Gly Gly Leu Glu Glu Arg Glu Ser Val 610 615 620 Gly Pro Leu Arg Glu Asp Phe Ser Leu Ser Ser Ser Ala Ser Ile Gly 625 630 635 640 Leu Leu Val Ile Ala Val Ala Ile Ala Thr Val Ile Val Ile Ser Leu 645 650 655 Val Met Leu Arg Lys Arg Gin Val Cys Thr Ile Ser His Gly Ile Val 660 665 670 Glu Val Asp Pro Met Leu Thr Pro Glu Glu Arg His Leu Asn Lys Met 675 680 685 Gin Asn His Gly Tyr Glu Asn Pro Thr Tyr Lys Thr Leu Glu Gin Met 690 695 700 Gin Ile 705 (2) INFORMATION FOR SEQ ID NO:30: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 695 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:30: Met Leu Pro Gly Leu Ala Leu Leu Leu Leu Ala Ala Trp Thr Ala Arg 1 5 10 15 Ala Leu Glu Val Pro Thr Asp Gly Asn Ala Gly Leu Leu Ala Glu Pro 20 25 30 Gin Ile Ala Met Phe Cys Gly Arg Leu Asn Met His Met Asn Val Gin 35 40 45 Asn Gly Lys Trp Asp Ser Asp Pro Ser Gly Thr Lys Thr Cys Ile Asp 50 55 60 Thr Lys Glu Gly Ile Leu Gin Tyr Cys Gin Glu Val Tyr Pro Glu Leu 65 70 75 80 Gin Ile Thr Asn Val Val Glu Ala Asn Gin Pro Val Thr Ile Gin Asn 85 90 95 96/15265 -88- Trp Cys Lys Arg Gly Arg Lys Gin Cys Lys Thr His Pro His Phe Val 100 105 110 Ile Pro Tyr Arg Cys Leu Val Gly Glu Phe Val Ser Asp Ala Leu Leu 115 120 125 Val Pro Asp Lys Cys Lys Phe Leu His Gin Glu Arg Met Asp Val Cys 130 135 140 Glu Thr His Leu His Trp His Thr Val Ala Lys Glu Thr Cys Ser Glu 145 150 155 160 Lys Ser Thr Asn Leu His Asp Tyr Gly Met Leu Leu Pro Cys Gly Ile 165 170 175 Asp Lys Phe Arg Gly Val Glu Phe Val Cys Cys Pro Leu Ala Glu Glu 180 185 190 Ser Asp Asn Val Asp Ser Ala Asp Ala Glu Glu Asp Asp Ser Asp Val 195 200 205 Trp Trp Gly Gly Ala Asp Thr Asp Tyr Ala Asp Gly Ser Glu Asp Lys 210 215 220 Val Val Glu Val Ala Glu Glu Glu Glu Val Ala Glu Val Glu Glu Glu 225 230 235 240 Glu Ala Asp Asp Asp Glu Asp Asp Glu Asp Gly Asp Glu Val Glu Glu 245 250 255 Glu Ala Glu Glu Pro Tyr Glu Glu Ala Thr Glu Arg Thr Thr Ser Ile 260 265 270 Ala Thr Thr Thr Thr Thr Thr Thr Glu Ser Val Glu Glu Val Val Arg 275 280 285 Val Pro Thr Thr Ala Ala Ser Thr Pro Asp Ala Val Asp Lys Tyr Leu 290 295 300 Glu Thr Pro Gly Asp Glu Asn Glu His Ala His Phe Gin Lys Ala Lys 305 310 315 320 Glu Arg Leu Glu Ala Lys His Arg Glu Arg Met Ser Gin Val Met Arg 325 330 335 Glu Trp Glu Glu Ala Glu Arg Gin Ala Lys Asn Leu Pro Lys Ala Asp 340 345 350 Lys Lys Ala Val Ile Gin His Phe Gin Glu Lys Val Glu Ser Leu Glu 355 360 365 Gin Glu Ala Ala Asn Glu Arg Gin Gin Leu Val Glu Thr His Met Ala 370 375 380 Arg Val Glu Ala Met Leu Asn Asp Arg Arg Arg Leu Ala Leu Glu Asn 385 390 395 400 Tyr Ile Thr Ala Leu Gin Ala Val Pro Pro Arg Pro Arg His Val Phe 405 410 415 Asn Met Leu Lys Lys Tyr Val Arg Ala Glu Gin Lys Asp Arg Gin His 420 425 430 Thr Leu Lys His Phe Glu His Val Arg Met Val Asp Pro Lys Lys Ala 435 440 445 Ala Gin Ile Arg Ser Gin Val Met Thr His Leu Arg Val Ile Tyr Glu 450 455 460 Arg Met Asn Gin Ser Leu Ser Leu Leu Tyr Asn Val Pro Ala Val Ala 465 470 475 480 Glu Glu Ile Gin Asp Glu Val Asp Glu Leu Leu Gin Lys Glu Gin Asn 485 490 495 Tyr Ser Asp Asp Val Leu Ala Asn Met Ile Ser Glu Pro Arg Ile Ser 500 505 510 Tyr Gly Asn Asp Ala Leu Met Pro Ser Leu Thr Glu Thr Lys Thr Thr 515 520 525 Val Glu Leu Leu Pro Val Asn Gly Glu Phe Ser Leu Asp Asp Leu Gin 530 535 540 Pro Trp His Ser Phe Gly Ala Asp Ser Val Pro Ala Asn Thr Glu Asn 545 550 555 560 Glu Val Glu Pro Val Asp Ala Arg Pro Ala Ala Asp Arg Gly Leu Thr 565 570 575 Thr Arg Pro Gly Ser Gly Leu Thr Asn Ile Lys Thr Glu Glu Ile Ser 580 585 590 Glu Val Lys Met Asp Ala Glu Phe Arg His Asp Ser Gly Tyr Glu Val 595 600 605 His His Gin Lys Leu Val Phe Phe Ala Glu Asp Val Gly Ser Asn Lys 610 615 620 Gly Ala Ile Ile Gly Leu Asn Val Gly Gly Val Val Ile Ala Thr Val 625 630 635 640 Ile Val Ile Thr Leu Val Met Leu Lys Lys Lys Gin Tyr Thr Ser Ile 645 650 655 His Glu Gly Val Val Glu Val Asp Ala Ala Val Thr Pro Glu Glu Arg 660 665 670 His Leu Ser Lys Met Gin Gin Asn Gly Tyr Glu Asn Pro Thr Thr Lys 675 680 685 Phe Phe Glu Gin Met Gin Asn 690 695 (2) INFORMATION FOR SEQ ID NO:31: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 507 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:31: Met Pro Leu Leu Leu Leu Leu Ala Trp Thr Ala Ala Val Gly Ala Leu 1 5 10 15 Ala Glu Pro Gin Ile Ala Met Phe Cys Gly Arg Leu Asn Met His Val 20 25 30 Asn Val Gin Gly Lys Trp Asp Ser Asp Pro Gly Thr Lys Cys Ile Thr 35 40 45 Lys Glu Gly Ile Leu Gin Tyr Cys Gin Glu Val Tyr Pro Glu Leu Gin 50 55 60 Ile Thr Asn Val Val Glu Ala Asn Gin Pro Val Ile Gin Asn Trp Cys 65 70 75 80 Lys Arg Asp Arg Lys Gin Cys Lys His Phe Val Ile Pro Tyr Arg Cys 85 90 95 Leu Val Gly Glu Phe Val Ser Asp Ala Leu Leu Val Pro Asp Lys Cys 100 105 110 Lys Phe Leu His Gin Glu Arg Met Asp Val Cys Glu His His Trp His 115 120 125 Thr Val Ala Lys Glu Cys Ser Ser Leu His Asp Tyr Gly Met Leu Leu 130 135 140 Pro Cys Gly Asp Asp Lys Phe Arg Gly Val Glu Phe Val Cys Cys Pro 145 150 155 160 Ala Glu Asp Val Ala Asp Glu Glu Asp Asp Asp Gly Gly Asp Asp Asp 165 170 175 Gly Glu Glu Ala Ile Glu Glu Val Glu Glu Asp Asp Asp Glu Asp Asp 180 185 190 96/15265 PCIYUS95/14416 -91- Glu Asp Gly Asp Glu Val Glu Glu Glu Glu Tyr Glu Glu Glu Arg Thr 195 200 205 Ala Thr Thr Thr Val Val Val Arg Val Pro Thr Ala Thr Asp Ala Val 210 215 220 Asp Tyr Leu Glu Thr Pro Gly Asp Glu Asn Glu His Ala His Phe Gin 225 230 235 240 Lys Ala Lys Glu Leu Glu Lys His Arg Glu Arg Met Gin Val Met Arg 245 250 255 Glu Trp Glu Glu Ala Glu Gin Ala Lys Asn Leu Pro Lys Ala Asp Lys 260 265 270 Ala Val Ile Gin His Phe Gin Val Glu Leu Glu Gin Glu Ala Ala Glu 275 280 285 Arg Gin Gin Leu Val Glu Thr His Met Ala Arg Val Glu Ala Met Leu 290 295 300 Asn Asp Arg Arg Leu Ala Leu Glu Asn Tyr Ile Ala Leu Gin Ala Pro 305 310 315 320 Pro Arg Pro Val Phe Leu Lys Lys Tyr Val Arg Ala Glu Gin Lys Asp 325 330 335 Arg His Thr Leu Lys His Phe Glu His Val Val Asp Pro Lys Ala Ala 340 345 350 Gin Ile Arg Ser Gin Val Met Thr His Leu Arg Val Ile Glu Arg Met 355 360 365 Asn Gin Ser Leu Ser Leu Leu Tyr Pro Val Ala Glu Glu Ile Asp Glu 370 375 380 Asp Glu Leu Leu Gin Glu Ala Asn Met Asp Leu Pro Ser Leu Glu Thr 385 390 395 400 Val Leu Asn Gly Glu Ser Leu Leu Gin Pro Trp His Phe Ala Asn Thr 405 410 415 Glu Glu Val Glu Asp Ala Ala Ala Leu Ser Gly Asn Ile Glu Glu Glu 420 425 430 Lys Met Ala Glu Lys Ser Gly Glu His Gin Leu Phe Glu Asp Gly Gly 435 440 445 Ala Ile Gly Leu Met Val Gly Val Ile Ala Thr Val Ile Val Ile Leu 450 455 460 Val Met Leu Lys Lys Lys Gin Tyr Ile His Gly Val Val Glu Val Asp 465 470 475 480 Ala Val Thr Pro Glu Glu Arg His Leu Lys Met Gin Asn Gly Tyr Glu 485 490 495 Asn Pro Thr Tyr Lys Phe Phe Glu Gin Met Gin 500 505 (2) INFORMATION FOR SEQ ID NO:32: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 532 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:32: Met Ala Ala Thr Gly Ala Ala Arg Leu Leu Leu Leu Leu Val Gly Leu 1 5 10 15 Thr Ala Ala Ala Ala Leu Ala Ala Leu Ala Ala Gly Ala Val Ala Glu 20 25 30 Pro Gin Ile Ala Met Phe Cys Gly Lys Leu Asn Met His Met Asn Ile 35 40 45 Gin Thr Gly Lys Trp Glu Pro Asp Pro Gly Thr Lys Cys Phe Thr Lys 50 55 60 Glu Glu Val Leu Gin Tyr Cys Gin Glu Met Tyr Pro Glu Leu Gin Ile 65 70 75 80 Thr Asn Val Met Glu Ala Asn Gin Val Ile Asp Asn Trp Cys Arg Arg 85 90 95 Gly Lys Lys Gin Cys Lys Arg Phe Val Pro Phe Lys Cys Leu Val Gly 100 105 110 Glu Phe Val Ser Asp Leu Leu Val Pro Glu Lys Cys Arg Phe Phe His 115 120 125 Glu Arg Met Glu Val Cys Glu His His Trp His Thr Val Lys Glu Ala 130 135 140 Cys Gin Gly Leu Ser Tyr Gly Met Leu Leu Pro Cys Gly Val Asp Phe 145 150 155 160 His Gly Glu Tyr Val Cys Cys Pro Asp Val Ser Glu Glu Glu Glu Asp 165 170 175 Glu Glu Glu Glu Glu Asp Glu Glu Glu Asp Asp Glu Pro Glu Asp Leu 180 185 190 Glu Asp Phe Glu Asp Asp Asp Glu Asp Glu Glu Glu Gly Glu Glu Val 195 200 205 Glu Asp Asp Tyr Asp Glu Glu Pro Thr Pro Gly Ser Thr Ile Val Lys 210 215 220 Val Pro Pro Thr Pro Pro Thr Asp Val Asp Val Tyr Phe Glu Thr Ala 225 230 235 240 Asp Asp Asn Glu His Ala Arg Phe Gin Lys Ala Lys Glu Gin Leu Glu 245 250 255 Arg His Arg Asn Arg Met Asp Val Lys Glu Trp Glu Glu Ala Glu Gin 260 265 270 Ala Lys Asn Leu Pro Lys Ala Glu Arg Gin Leu Ile Gin His Phe Gin 275 280 285 Met Val Leu Glu Glu Ala Ala Ser Glu Gin Gin Leu Val Glu Thr His 290 295 300 Leu Ala Arg Val Glu Ala Met Leu Asn Asp Arg Arg Met Ala Leu Glu 305 310 315 320 Asn Tyr Leu Ala Ala Leu Gin Ser Asp Pro Pro Arg Pro Arg Ile Leu 325 330 335 Pro Leu Arg Arg Tyr Val Arg Ala Glu Lys Asp Arg His Thr Ile Arg 340 345 350 His Tyr Gin His Val Ala Val Asp Pro Glu Lys Ala Ala Gin Met Lys 355 360 365 Ser Gin Val Met Thr His Leu His Val Ile Glu Glu Arg Asn Gin Ser 370 375 380 Leu Ser Leu Leu Tyr Asp Pro Val Ala Gin Glu Asp Glu Leu Leu Gin 385 390 395 400 Ala Glu Arg Ala Asp Met Asp Phe Ala Ser Ile Ser Glu Thr Val Val 405 410 415 Ser Ser Glu Ser Glu Ile Pro Phe His Pro Phe His Pro Phe Pro Ala 420 425 430 Asp Thr Gin Glu Leu Glu Gly Gly Gly Ala Val Ser Glu Asn Met Asp 435 440 445 Glu Asp Met Asn Ala Arg Gly Gly Leu Glu Glu Arg Glu Val Gly Leu 450 455 460 Glu Asp Ser Ser Ser Ala Ile Gly Leu Leu Val Ile Ala Val Ala Ile 465 470 475 480 Ala Thr Val Ile Val Ile Ser Leu Val Met Leu Arg Lys Arg Gin Thr 485 490 495 Ile Ser His Gly Ile Val Glu Val Asp Pro Met Leu Thr Pro Glu Glu 500 505 510 Arg His Leu Lys Met Gin His Gly Tyr Glu Asn Pro Thr Tyr Lys Leu 515 520 525 Glu Gin Met Gin 530 (2) INFORMATION FOR SEQ ID NO:33: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 429 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (xi) SEQUENCE DESCRIPTION: SEQ ID NO:33: Met Gly Pro Thr Ser Ala Ala Arg Pro Leu Leu Leu Leu Leu Ser Leu 1 5 10 15 Ala Leu Ala Ala Val Gly Pro Ala Ala Gly Ala Gin Val Ala Leu Cys 20 25 30 Gly Arg Leu Leu His Asp Leu Thr Gly Arg Trp Glu Pro Asp Pro Ser 35 40 45 Arg Cys Leu Gin Val Leu Glu Tyr Cys Gin Met Tyr Pro Glu Leu His 50 55 60 Ile Val Gin Ala Gin Ala Ile Met Glu Trp Cys Arg Cys His Val Val 65 70 75 80 Pro Phe Cys Leu Pro Gly Glu Phe Val Ser Glu Ala Leu Leu Val Pro 85 90 95 Glu Gly Cys Arg Phe Leu His Gin Glu Arg Met Asp Cys Glu Arg Arg 100 105 110 His Ala Glu Ala Cys Ser Ser Gin Gly Leu His Gly Gly Met Leu Leu 115 120 125 Pro Cys Gly Asp Arg Phe Arg Gly Val Glu Tyr Val Cys Cys Pro Pro 130 135 140 Asn Gly Ala Asp Gly Gly Glu Gly Gly Glu Asp Glu Glu Glu Val Glu 145 150 155 160 Phe Gin Asp Glu Gin Glu Glu Glu Glu Glu Glu Glu Arg Ala Pro Ser 165 170 175 Thr Val Arg Val Pro Thr Pro Pro Thr Asp Gly Val Asp Val Tyr Phe 180 185 190 Gly Pro Gly Glu Glu His Phe Arg Ala Lys Asp Leu Glu Glu Arg Arg 195 200 205 Ile Asn Glu Val Met Arg Glu Trp Ala Asp Gin Lys Asn Leu Pro Lys 210 215 220 Ala Asp Arg Gin Ala Leu Glu His Phe Gin Ile Leu Gin Leu Glu Glu 225 230 235 240 Glu Gin Gly Glu Arg Gin Leu Val Glu Thr His Arg Val Ala Leu Ile 245 250 255 Asn Asp Arg Arg Ala Leu Glu Phe Leu Ala Ala Leu Gin Gly Asp Pro 260 265 270 Pro Ala Arg Val Leu Ala Leu Arg Arg Tyr Leu Arg Ala Glu Gin Lys 275 280 285 Glu His Thr Leu Arg His Tyr Gin His Val Ala Val Asp Pro Glu Lys 290 295 300 Ala Gin Met Arg Gin Val Thr His Leu Val Ile Glu Glu Arg Met Asn 305 310 315 320 Gin Ser Leu Gly Leu Leu Asn Pro Leu Ala Gin Glu Leu Gin Gin Glu 325 330 335 Leu Leu Ala Glu His Ser Glu Val Ser Ser Glu Ser Leu Pro Asp Pro 340 345 350 Pro Thr Glu Ser Ser Gly Glu Glu Glu Lys Ala Ala Arg Phe Asp Gin 355 360 365 Arg Asp Leu Ala Gly Gly Ser Ala Gly Leu Leu Ile Met Gly Gly Gly 370 375 380 Leu Ile Val Leu Ser Leu Leu Leu Leu Arg Lys Lys Tyr Thr Ile Ser 385 390 395 400 His Gly Val Val Glu Val Asp Pro Met Leu Thr Glu Glu Gin Leu Leu 405 410 415 Gin His Gly Tyr Glu Asn Pro Thr Tyr Phe Leu Glu Glu 420 425

Claims: What Is Claimed Is: 1. A method for modulating transcription from the amyloid β- protein precursor (APP) promoter, comprising: binding die APP promoter with a transcriptional activator selected from recombinant upstream stimulatory factor (rUSF) or a composition comprising native USF whereby transcription from the APP promoter is activated. 2. The method of claim 1, wherein the APP promoter is operably linked to a nucleic acid sequence encoding APP. 3. The method of claim 1, wherein the APP promoter is operably linked to a nucleic acid sequence encoding a reporter protein. 4. The method of claim 3, wherein said reporter protein is the luciferase reporter protein. 5. The method of claim 1, wherein transcription from the APP promoter is detected by primer extension analysis of the resulting transcript. 6. The method of claim 1 , wherein transcription is activated by transfecting a cell containing the APP promoter with a DNA or RNA construct which is capable of expressing USF. 7. The method of claim 1, wherein said transcriptional activator binds to die AP-l/AP-4 site present in the APP promoter. 8. The method of claim 1 , further comprising: contacting said transcriptional activator with a USF binding compound which is capable of down-regulating transcription from the APP promoter. 9. The method of claim 8, wherein said USF binding compound is selected from APLP, APP, or a nucleic acid sequence comprising the USF consensus sequence. 10. The method of claim 9, wherein said USF binding compound is an APLP selected from APLPl or APLP2. 11. A method for down-regulating transcription from the amyloid β- protein precursor (APP) promoter, comprising: contacting USF with a USF binding compound capable of interfering with USF binding to d e APP promoter whereby transcription from the APP promoter is down-regulated. 12. The method of claim 11, wherein said USF binding compound is selected from an APLP, APP, or a nucleic acid sequence comprising the USF consensus sequence. 13. The method of claim 12, wherein said USF binding compound is an APLP selected from APLP-1 or APLP-2. 14. The method of claim 13 , wherein transcription is down-regulated by transfecting a cell containing the APP promoter with a DNA or RNA construct which is capable of expressing said APLP protein. 15. The metiiod of claim 11 , wherein die APP promoter is operably linked to a nucleic acid sequence encoding APP. 16. The metiiod of claim 11 , wherein the APP promoter is operably linked to a reporter protein. 17. A method for screening candidate upstream stimulatory factor (USF) binding compounds to determine which are capable of down-regulating transcription from the amyloid β-protein precursor (APP) promoter, comprising the steps of: (a) transfecting a host cell with a DNA or RNA construct containing the APP promoter operably linked to a gene encoding a reporter protein; (b) transfecting said host cell widi a DNA or RNA construct capable of expressing upstream stimulatory factor (USF) protein; (c) measuring reporter protein expression activated by USF binding to die APP promoter; (d) transfecting said host cell widi a DNA or RNA construct either containing or capable of expressing a USF binding compound; and (e) measuring if a decrease in reporter protein expression occurs due to said USF binding compound interfering with USF binding to die APP promoter. 18. The method of claim 17, wherein said gene encoding a reporter protein encodes a luciferase reporter protein. 19. The method of claim 17, wherein said DNA binding compound is an APLP. 20. The method of claim 19, wherein said APLP is selected from APLP-1 or APLP-2.

Inventors: KOVACS DORA M, TANZI RUDOLPH E, KOVACS, DORA, M., TANZI, RUDOLPH, E.
Applicants: GEN HOSPITAL CORP, THE GENERAL HOSPITAL CORPORATION
Application year: 1995
Main classification: C12Q1
IPC codes: A61K38, C07K14, C12N5, C12N15, C12Q1