bluuebunny/crossref_metadata_2025 huggingface dataset:
Alterations in gene expression in sorghum induced by the host-specific toxin from
Periconia circinata
Authors: Wolpert Thomas J. , Dunkle Larry D.
Published: May 2006

Susceptibility of sorghum to the fungal pathogen
Periconia circinata
and sensitivity to its host-specific toxin are determined by the semidominant allele at the
pc
locus. Pretreatment of susceptible seedlings with cycloheximide or cordycepin for 4 hr before treatment with the toxin protected the seedlings against toxin-induced loss of electrolytes and prevented development of disease symptoms.
In vivo
incorporation of [
3
H]leucine into protein was inhibited 91% and 47% by cycloheximide and cordycepin, respectively, but was not affected by the toxin. Gel electrophoresis and fluorography of
in vivo
-labeled proteins extracted from non-treated and toxin-treated root tips of near-isogenic susceptible and resistant lines revealed a selective increase in radioactivity of a protein band at
M
r
16,000 only in preparations from toxin-treated susceptible root tips. Two-dimensional gel electrophoresis separated the
M
r
16,000 band into four proteins and confirmed the increased rate of synthesis. Products of
in vitro
translation were substantially enriched with the four
M
r
16,000 proteins when total RNA from toxin-treated susceptible root tips was used in a cell-free protein-synthesizing system. Because the proteins that increase are common to both susceptible and resistant genotypes, the toxin apparently interferes with a regulatory function, perhaps a function of the
pc
locus, and thereby alters gene expression in the susceptible genotype. The data suggest but do not establish that phytotoxicity results from the increased rate of synthesis of the specific proteins.
Localization of cellular retinol-binding protein in several rat tissues.
Authors: Porter S B , Fraker L D , Chytil F , Ong D E
Published: May 2006

The distribution of cellular retinol-binding protein (CRBP) in rat liver, ileum, and epididymis was examined by the peroxidase-antiperoxidase immunolocalization technique. Positive cytoplasmic staining was seen in the liver when antiserum prepared against purified CRBP was used but not when antiserum absorbed with purified CRBP was used. Ileal mucosa, a tissue that contains no detectable CRBP, showed no positive staining. The epididymis showed strong positive staining in the caput but not in the cauda. Staining was present in principal and basal cells but not in peritubular or interstitial cells. Radioimmunoassay revealed that the CRBP within the caput epididymidis was localized in the initial segment and proximal region, areas known to be involved in the synthesis and secretion of factors necessary for sperm maturation. The results demonstrate that the expression of CRBP may vary within the same cell type, as well as between different cell types within the same tissue.
Enzymatic deacylation of the lipid A moiety of Salmonella typhimurium lipopolysaccharides by human neutrophils.
Authors: Hall C L , Munford R S
Published: May 2006

Lipid A, the toxic moiety of Gram-negative bacterial lipopolysaccharides (endotoxins), is a glucosamine disaccharide to which fatty acid and phosphate residues are covalently attached. Recent studies of Salmonella lipid A indicate that 3-hydroxytetradecanoic acid (3-OH-14:0) residues are directly linked to the glucosamine backbone and the nonhydroxylated fatty acids (principally dodecanoic and tetradecanoic acids) are esterified to the hydroxyl groups of some of the 3-OH-14:0 molecules. We report here that the granule fraction of human neutrophils contains one or more enzymes that partially deacylate Salmonella typhimurium lipid A by removing the nonhydroxylated fatty acids, leaving almost all of the 3-OH-14:0 residues linked to glucosamine. The available evidence suggests that similar reactions also occur in living neutrophils that ingest lipopolysaccharides by antibody-dependent phagocytosis.
Immunological crossreactivity of eukaryotic C1-tetrahydrofolate synthase and prokaryotic 10-formyltetrahydrofolate synthetase.
Authors: Staben C , Rabinowitz J C
Published: May 2006

Antiserum to yeast C1-tetrahydrofolate (C1-H4folate) synthase reacts with other eukaryotic C1-H4folate synthases and prokaryotic 10-formyltetrahydrofolate (10-CHO-H4folate) synthetases [formate:tetrahydrofolate ligase (ADP-forming), EC 6.3.4.3] even though these enzymes vary in subunit size and function and probably vary widely in sequence. The comigration of the purified enzymes with the immunoreactive material establishes the specificity of the reaction for C1-H4folate synthase proteins. Reciprocal crossreaction of the antibody to Clostridium acidiurici 10-CHO-H4folate synthetase with the eukaryotic proteins indicates that such broad cross-species reactions are not specific to the antisera elicited in response to the yeast C1-H4folate synthase. These specific crossreactions among divergent species have been observed only on an electrophoretic transfer blot of a denaturing polyacrylamide gel. These observations may have been possible because of the sensitivity and specificity of the technique, which differ from more conventional immunochemical methods.
Biosynthesis of renin: multiplicity of active and intermediate forms.
Authors: Pratt R E , Ouellette A J , Dzau V J
Published: May 2006

Processing of renin involves sequential proteolytic cleavages of a preproform to the active mature forms. Preprorenin is rapidly internalized cotranslationally into the rough endoplasmic reticulum and hydrolyzed by signal peptidase to produce prorenin. In the Golgi, prorenin is converted (within 15 min) to a form of renin that is enzymatically active. Over the next 12 hr, a slow intracellular process removes a dipeptide near the carboxyl terminus, converting the one-chain renin into two chains joined by a single disulfide bond. This conversion occurs during formation, condensation, and packaging of renin granules. The resultant two-chain renin is approximately one-sixth as active as the one-chain form. The intact renin molecule is obligatory for enzymatic activity because heavy chain alone has little or no activity. Both one- and two-chain renins are secreted, but prorenin is not. Multiple isoelectric forms of prorenin, one-chain renin, and two-chain renin are also observed. This microheterogeneity probably results from minor differences in amino acid composition as a consequence of variations in cleavage positions during processing. Thus, these data suggest that renin synthesis and secretion is complex and may be subject to regulation at multiple steps. Furthermore, based on the results of this study, we also propose that renin can be secreted by two different pathways.
Thermodynamics of assembly of Escherichia coli aspartate transcarbamoylase.
Authors: McCarthy M P , Allewell N M
Published: May 2006

Reaction microcalorimetry and potentiometry have been used to define the thermodynamics of assembly of Escherichia coli aspartate transcarbamoylase (aspartate carbamoyltransferase, carbamoylphosphate:L-aspartate carbamoyltransferase, EC 2.1.3.2) from its catalytic and regulatory subunits and the linkage between assembly and proton binding. Over the pH range 7-9.5 and the temperature range 15-30 degrees C, assembly is characterized by negative enthalpy and heat capacity changes and positive entropy changes. The dependence of the enthalpy and entropy changes on pH is complex; however, the negative heat capacity change results in both quantities becoming more negative with increasing temperature. Assembly is linked to the binding of protons; the effects observed can be fit to models involving six or more ionizable groups with pK values of 7.3-7.4, 8.5-8.8, and 9.2-9.5, which ionize cooperatively. Contributions from additional groups cannot be ruled out and are in fact expected. The overall pattern of thermodynamic effects implies a complex set of intersubunit interactions. Protonation reactions and increased hydrogen bonding are likely to be the major sources of the negative enthalpy change; however, the negative heat capacity change results primarily from changes in solvent structure associated with hydrophobic and electrostatic bond formation with changes in low-frequency vibrational modes making a secondary contribution. Similarly, the relatively small entropy change observed within the temperature range examined probably reflects the balance between positive contributions from increased hydrophobic and electrostatic bonding and negative contributions from increased hydrogen bonding and damping of low-frequency vibrational modes.
Autogenous production of a hemopoietic growth factor, persisting-cell-stimulating factor, as a mechanism for transformation of bone marrow-derived cells.
Authors: Schrader J W , Crapper R M
Published: May 2006

Lines of hemopoietic progenitor cells can be grown for prolonged or indefinite periods in vitro in the presence of a specific growth factor produced by activated T lymphocytes, persisting (P)-cell-stimulating factor (PSF). From such a PSF-dependent line, we report the emergence of variants that had concomitantly acquired both the capacity for autonomous growth in the absence of exogenous PSF and the capacity for the autogenous production of PSF. The link between these two new properties was strengthened by the demonstration that the variant lines absorbed PSF and, in some culture conditions, responded to exogenous PSF. Thus when variant cells were plated at low density in 1-ml agar cultures, cloning efficiency and colony size were enhanced by supplementation with sources of PSF, including medium conditioned by concanavalin A-stimulated normal spleen cells, T-cell tumors and T-cell hybridomas, and, importantly, medium conditioned by the autonomous P-cell lines themselves. In contrast to the parental line, the autonomous clones tested formed progressively growing tumors in vivo. It is proposed that the myelomonocytic leukemia WEHI-3B that produces PSF arose from a neutrophil-macrophage progenitor through acquisition of the capacity for the autogenous production of PSF and that the autogenous production of PSF may play a similar role in a range of proliferative disorders both of the blood and of other tissues containing components derived from the bone marrow.
Transcription of the
white
locus in
Drosophila melanogaster
Authors: O'Hare Kevin , Levis Robert , Rubin Gerald M.
Published: May 2006

Genetic studies of the
white
locus have shown that it has a distal region where structural mutations occur and a proximal region where regulatory mutations occur. To better understand the molecular basis of this genetic organization we have analyzed
white
locus transcription. A 2.7-kilobase transcript comprising 0.0005% of poly(A)-RNA was detected in RNA prepared from pupae or adults. The structure of this transcript helps clarify some unusual genetic properties of the locus. There is a small 5′ exon separated from the majority of the sequences found in the mature RNA by an intron of ≈2.8 kilobases. This 5′ exon is from the proximal region of the locus, whereas the main body of the RNA maps to the distal region. The mutationally silent region between the proximal and distal regions corresponds to the large intron. We have identified the family and determined the exact location of a number of transposable element insertions within the locus. These results show that transposable element insertions within introns can be without phenotypic effect. We have also investigated the effect on the
white
transcript of the
zeste
mutation, which represses
white
locus expression as judged by eye color phenotype. The RNA was unchanged in size or abundance in poly(A)-RNA from adult flies. This demonstrates that the
zeste-white
interaction does not occur by simply repressing transcription of the
white
locus in all tissues.
High-resolution chromosomal localization of human genes for amylase, proopiomelanocortin, somatostatin, and a DNA fragment (D3S1) by in situ hybridization.
Authors: Zabel B U , Naylor S L , Sakaguchi A Y , Bell G I , Shows T B
Published: May 2006

The method of in situ hybridization has become a significant technique for specific-site chromosome mapping. We show that the resolution of in situ hybridization can be increased by hybridizing the probe to stretched prometaphase chromosomes with high-resolution banding obtained after 5-bromodeoxyuridine treatment of the cells and with a Hoechst 33258/Giemsa chromosome-staining method. Using this procedure, we assigned to specific chromosome sites three cloned genes and one DNA polymorphism: amylase gene (AMY) to 1p21; proopiomelanocortin gene (POMC) to 2p23, somatostatin gene (SST) to 3q28, and a single copy DNA segment (D3S1) to 3q12.
Molecular map of the murine S region.
Authors: Chaplin D D , Woods D E , Whitehead A S , Goldberger G , Colten H R , Seidman J G
Published: May 2006

Eighteen overlapping cosmid clones spanning 240 kilobases and encoding the gene for factor B and two genes related to the fourth component of complement (C4) were isolated from a murine H-2d genomic library. Cosmid clones were identified by hybridization to human cDNA probes for factor B and C4 and were linked by chromosomal walking procedures. The cluster of clones contains two regions with sequences homologous to the C4 cDNA probe, both in the same orientation, representing a direct duplication of at least 55 kilobases of chromosomal DNA, separated by a shorter (less than 25 kilobases) segment of nonduplicated DNA. Restriction fragment-length polymorphism seen by using C4 probes maps these sequences to the S region of the major histocompatibility complex. 5' to the two C4-like sequences is an approximately equal to 40-kilobase-long region of chromosomal DNA remarkable for its lack of restriction fragment-length polymorphism, containing sequences homologous to the human factor B cDNA probe. These experiments demonstrate that the structural gene for factor B is located in the S region of the murine major histocompatibility complex and that this region contains an extensive direct duplication that contains the structural gene for mouse C4 and, we presume, for the sex-limited protein variant, Slp. RNA transfer blot analysis of total liver RNA from high C4- and low C4-producing strains showed that steady-state levels of C4-hybridizing RNA were much greater in high C4-producing strains. Regulation of circulating C4 levels in high C4 and low C4 strains is at least partly at the level of mRNA transcription, processing, or degradation.
Genetic analysis of the folding pathway for the tail spike protein of phage P22.
Authors: Goldenberg D P , Smith D H , King J
Published: May 2006

Temperature-sensitive mutations in the gene encoding the trimeric tail spike protein of phage P22 interfere with protein maturation at 39 degrees C. We show here that temperature-sensitive mutations at many sites block the folding pathway prior to accumulation of the partially folded protrimer intermediate. Temperature-shift experiments indicate that at least some of the mutants accumulate an earlier intermediate in the folding pathway. Immunoprecipitation experiments suggest that the conformation of the isolated temperature-sensitive polypeptide chains is closer to that of the unfolded chain than to that of the mature spike formed at permissive temperature. The sites of these mutations probably represent amino acid sequences that play key roles during the folding of the tail spike polypeptide chain but are not important in the mature protein.
Activation of the c-mos oncogene in a mouse plasmacytoma by insertion of an endogenous intracisternal A-particle genome.
Authors: Canaani E , Dreazen O , Klar A , Rechavi G , Ram D , Cohen J B , Givol D
Published: May 2006

The activation of the cellular oncogene c-mos in mouse plasmacytoma XRPC24 was found to result from the insertion of a 4.7-kilobase-pair cellular DNA element, within the c-mos coding region. The element terminates on both sides with a direct repeat of around 335 nucleotides. The repeat as well as internal sequences of the element show strong homology to endogenous intracisternal A-particle (IAP) genes. The IAP genome integrated within c-mos in a head-to-head (5' to 5') configuration. This juxtapositioned the IAP 5' long terminal repeat next to the bulk of the oncogene's coding region and shifted c-mos 5' coding and flanking sequences to a position further upstream. The significance of several aspects of this activation and transposition event is discussed.
Synthetic leader peptide modulates secretion of proteins from microinjected Xenopus oocytes.
Authors: Koren R , Burstein Y , Soreq H
Published: May 2006

To investigate the role of the leader peptide in modulating secretion from living cells, we injected a synthetic peptide into Xenopus oocytes. The peptide consisted of the NH2-terminal leader sequence of mouse immunoglobulin light chain precursor. We found that the leader peptide has two different roles in regulating secretion from the oocytes. First, it competitively inhibits the synthesis of secretory and membrane proteins but not of cytoplasmic proteins. The inhibition occurs both with oocyte proteins and with proteins directed by coinjected myeloma mRNA. The inhibition reaches a maximum 2 hr after injection and decays within 3 hr. It appears to be mediated through the cell membrane, because 125I-labeled leader peptide segregates into the membrane fraction of microinjected oocytes simultaneously with the interference with methionine incorporation. A second role of the microinjected leader peptide is to induce a rapid acceleration in the rate of export of secretory proteins from the oocyte. The maximal enhancement effect is obtained upon injection of 50 ng of leader peptide per oocyte. It is not merely due to the small size, negative charge, or hydrophobicity of the peptide, because enhanced secretion does not occur when glucagon, poly-L-glutamic acid, or Triton X-100 is injected. Furthermore, immunoreaction of the peptide with specific antibodies prior to microinjection prevents the accelerated export. Our observations indicate that in Xenopus oocytes, the leader peptide is involved in both translocation and later step(s) in the secretory pathway.
Polarization of the Golgi apparatus and the microtubule-organizing center within cloned natural killer cells bound to their targets.
Authors: Kupfer A , Dennert G , Singer S J
Published: May 2006

In cell couples formed between a cloned murine natural killer (NK) cell and either of two target cells, we have obtained evidence by immunofluorescence observations for a rapid coordinate repositioning of two organelles, the microtubule organizing center and the perinuclear Golgi apparatus, inside the NK cell to face the region of contact with the target cell. With microtubule-disrupting drugs, which also cause a dispersion of the Golgi apparatus throughout the periphery of the NK cell, it was found that target cell lysis is reversibly inhibited. It is proposed that the coordinate repositioning of the two organelles serves the function of directing secretory vesicles derived from the Golgi apparatus to the bound target cells, the secretory vesicles containing components that induce target cell lysis.
Exon-intron organization and complete nucleotide sequence of a human major histocompatibility antigen DC beta gene.
Authors: Larhammar D , Hyldig-Nielsen J J , Servenius B , Andersson G , Rask L , Peterson P A
Published: May 2006

We have determined the complete nucleotide sequence of a human class II histocompatibility antigen DC beta gene. The gene spans more than 7 kilobases and contains five exons corresponding to the different domains of the DC beta polypeptide. The exon-intron organization is thus analogous to that of class II antigen alpha-chain genes, class I antigen heavy chain genes, and the constant parts of immunoglobulin genes, emphasizing further the evolutionary relationship among these molecules. The mature polypeptide deduced from the DC beta gene shows 93% and 88% homology, respectively, to sequences derived from two DC beta cDNA clones of other haplotypes. The allelic polymorphism of DC beta chains resides predominantly in the first extracellular domain, whereas the rest of the polypeptide is virtually constant. The exons of the DC beta gene display high homology to the corresponding exons of a murine I-A beta gene. Also, the introns show significant homology. The DC beta chains lack eight amino acids in the cytoplasmic tail, as compared to DR and I-A beta chains. This is probably due to a nonfunctional splice junction of DC beta genes, causing a separate cytoplasmic exon to be nonexpressed.
Identification of cDNA clones encoding secretory isoenzyme forms: sequence determination of canine pancreatic prechymotrypsinogen 2 mRNA.
Authors: Pinsky S D , LaForge K S , Luc V , Scheele G
Published: May 2006

A cDNA library has been constructed from canine poly(A)+ mRNA. Clones containing cDNA inserts coding for prechymotrypsinogen 2 (isoelectric point = 7.1; Mr = 27,500), one of three canine pancreatic isoenzyme forms, were selected by colony hybridization using a cDNA probe synthesized from immunoselected prechymotrypsinogen 2 mRNA. To verify that cDNA clones code for prechymotrypsinogen 2 forms that translocate across rough endoplasmic reticulum membranes and fold into stable and identifiable secretory proteins, we conducted in vitro translation of hybrid-selected mRNA in the presence of microsomal membranes and optimal concentrations of glutathione and analyzed nascent translation products in their nonreduced state by two-dimensional isoelectric focusing/NaDodSO4 gel electrophoresis and fluorography. A near full-length chymotrypsinogen 2 cDNA and its primed extension were used to determine the nucleotide sequence for the entire coding region of prechymotrypsinogen 2 mRNA and 87 residues, including a poly(A) addition signal, in the 3' nontranslated region. The deduced amino acid sequence shows a 263-residue presecretory protein containing an 18-residue amino-terminal transport peptide (Met-Ala-Phe-Leu-Trp-Leu-Leu-Ser-Cys-Phe-Ala-Leu-Leu-Gly-Thr-Ala-Phe-Gly ), which we have previously shown to mediate the translocation of chymotrypsinogen 2 across the rough endoplasmic reticulum membrane. Following the transport peptide is a 245-residue proenzyme, which shows 82% and 80% sequence identity with bovine chymotrypsinogens A and B, respectively. Conserved among the three zymogens are 10 Cys residues that form five disulfide bonds in bovine chymotrypsinogens A and B and the residues that are required for zymogen activation, substrate binding, and catalytic activity.
Translocation of an immunoglobulin kappa locus to a region 3' of an unrearranged c-myc oncogene enhances c-myc transcription.
Authors: Erikson J , Nishikura K , ar-Rushdi A , Finan J , Emanuel B , Lenoir G , Nowell P C , Croce C M
Published: May 2006

We have studied somatic cell hybrids between mouse myeloma and JI Burkitt lymphoma cells carrying a t(2;8) chromosome translocation for the expression of human kappa chains. and for the presence and rearrangements of the human c-myc oncogene and kappa chain genes. Our results indicate that the c-myc oncogene is unrearranged and remains on the 8q+ chromosome of JI cells. Two rearranged C kappa genes were detected: the expressed allele on normal chromosome 2 and the excluded kappa allele that was translocated from chromosome 2 to the involved chromosome 8 (8q+). The distribution of V kappa and C kappa genes in hybrid clones retaining different human chromosomes indicated that C kappa is distal to V kappa on 2p and that the breakpoint in this Burkitt lymphoma is within the region carrying V kappa genes. High levels of transcripts of the c-myc gene were found when it resided on the 8q+ chromosome but not on the normal chromosome 8, demonstrating that translocation of a kappa locus to region distal to the c-myc oncogene enhances c-myc transcription.
Enveloped double-stranded DNA insect virus with novel structure and cytopathology
Authors: Federici Brian A.
Published: May 2006

An unusual type of virus has been isolated from larvae of the cabbage looper,
Trichoplusia ni
(Lepidoptera; Noctuidae). The virus infects a variety of tissues, including fat body, epidermis, and tracheal matrix, causing a chronic, fatal disease. Viral replication begins in the nucleus and is accompanied by invagination of the nuclear envelope and extensive nuclear and cellular hypertrophy. The nuclear envelope eventually ruptures and fragments, after which viral-induced membranes are assembled along planes through the cell and around its periphery. Subsequently, these membranes coalesce, partitioning most of the cell, including viroplasms and virions in various stages of assembly, among a cluster of vesicles. The vesicles dissociate and are liberated into the hemolymph where they accumulate in large numbers (>10
8
vesicles per ml), causing the blood to become opaque white. The virus has been isolated from
T. ni
and transmitted
per os
and by injection to
T. ni
and several other species of the family Noctuidae. The virions produced by this virus are large (
ca.
130 × 400 nm), enveloped, and allantoid in shape with complex symmetry and contain apparently linear, double-stranded DNA of
M
r
of ≈ 1.00 × 10
8
. The envelope contains subunits arranged in a hexagonal pattern that impart a distinctive reticular appearance to virions in negatively stained preparations. The unique structural and developmental properties of this virus indicate that it is a member of a group of enveloped, double-stranded DNA viruses not observed previously.
N-acetylaspartylglutamate: an endogenous peptide with high affinity for a brain "glutamate" receptor.
Authors: Zaczek R , Koller K , Cotter R , Heller D , Coyle J T
Published: May 2006

A brain peptide with high affinity (420 nM) and marked specificity for brain receptor sites labeled with L-[3H]glutamate has been identified. Amino acid analysis and mass spectroscopy indicate that the peptide is N-acetylaspartylglutamate. The peptide exhibits potent convulsant properties when injected into the rat hippocampus, similar to those produced by the glutamate receptor agonist, quisqualic acid. These findings raise the question whether endogenous brain peptides enriched in acidic amino acids may serve as excitatory neurotransmitters.
At what wavelengths should we search for signals from extraterrestrial intelligence?
Authors: Townes C. H.
Published: May 2006

It has often been concluded that searches for extraterrestrial intelligence (SETI) should concentrate on attempts to receive signals in the microwave region, the argument being given that communication can occur there at minimum broadcasted power. Such a conclusion is shown to result only under a restricted set of assumptions. If generalized types of detection are considered—in particular, photon detection rather than linear detection alone—and if advantage is taken of the directivity of telescopes at short wavelengths, then somewhat less power is required for communication at infrared wavelengths than in the microwave region. Furthermore, a variety of parameters other than power alone may be chosen for optimization by an extraterrestrial civilization. Hence, while partially satisfying arguments may be given about optimal wavelengths for a search for signals from extraterrestrial intelligence, considerable uncertainty must remain.
Endogenous inhibitors of lysosomal proteinases.
Authors: Pontremoli S , Melloni E , Salamino F , Sparatore B , Michetti M , Horecker B L
Published: May 2006

Specific inhibitors of three lysosomal proteinases are present in the cytosolic and lysosomal compartments of rabbit liver. The cytosolic inhibitors, purified by chromatography on DEAE-Trisacryl and Sephadex G-75, show specificities toward cathepsin M, cathepsins B and L, and fructose 1,6-bisphosphatase converting enzyme (CE), respectively, and are designated IM, IB/L, and ICE. Inhibitors with similar specificities have been isolated from the intralysosomal compartment. Two of these inhibitors, IM and ICE, are also present in the lysosomal membranes. The lysosomal distribution parallels that of the respective proteinases. The inhibitors are polypeptides with molecular weights of 5,000-10,000 for the two forms of IB/L, 12,500 for IM, and 10,000-40,000 for the ICE species.
Band patterns seen by electron microscopy in ordered arrays of bovine and human fibrinogen and fibrin after negative staining.
Authors: Williams R C
Published: May 2006

When fibers of fibrin clots or fragments of fibrinogen pellets are negatively stained they exhibit in the electron microscope characteristic patterns of cross-striations or bands. Those found in pellet material are indistinguishable from those seen in thrombin-induced fibrin fibers. The pattern seen in fibrin from bovine sources contains three equally spaced faint bands between every two of the broad prominent ones, spaced 23 nm apart. Human material shows a different pattern, one wherein no central faint band is seen, whereas the two remaining ones are broader. Its character is unaffected by crosslinking following fiber formation and preceding negative staining. The bovine pattern, however, is converted by such crosslinking to one that closely resembles the human. It is suggested that the striation pattern in human fibrin is due to juxtapositions of E domains of the parallel-aligned fibrin monomers with tightly coiled COOH-terminal regions of beta and gamma polypeptide chains, with no discernible contribution to the pattern from the alpha chain. In negatively stained untreated fibers of bovine fibrin, however, it is proposed that the COOH-terminal region of the alpha chain becomes tightly coiled, thereby contributing the faint central striation to the band pattern. Crosslinking prevents this conformational change in the alpha chain.
Polymorphic restriction endonuclease fragment segregates and correlates with the gene for HLA-B8.
Authors: Cann H M , Ascanio L , Paul P , Marcadet A , Dausset J , Cohen D
Published: May 2006

Cellular DNA from HLA-typed individuals was digested with the restriction endonuclease EcoRV. After electrophoresis and transfer to a hybridization membrane, the restriction endonuclease fragments were probed with cDNA carrying the nucleotide sequence encoding a class 1 HLA gene. Polymorphism for presence or absence of various EcoRV fragments was noted in a panel of unrelated HLA-typed individuals. A polymorphic 8.6-kilobase pair EcoRV fragment was found which correlated in the panel with the serologically defined gene HLA-B8. A family study revealed that this fragment segregated with the haplotype carrying the HLA-B8 gene. This fragment may carry the gene for HLA-B8 or it may represent another class 1 gene (or pseudogene) in linkage disequilibrium with HLA-B8.
Functional changes of intermediate filaments in fibroblastic cells revealed by a monoclonal antibody.
Authors: Dulbecco R , Allen R , Okada S , Bowman M
Published: May 2006

We describe reversible changes of intermediate filaments of fibroblastic cells associated with changes in the functional state of the cells. The changes are revealed by comparing the immunofluorescence patterns given by a monoclonal antibody and a polyclonal serum, both recognizing vimentin. The state of the filaments depends on culture density; this effect cannot be attributed to the nutritional state of the cells, their growth rate, or substances released into the medium. It seems to depend mainly on the aggregation of filaments during strong cell movements. The possible significance of these findings for the functional role of intermediate filaments is discussed.
Cellular myc oncogene is altered by chromosome translocation to an immunoglobulin locus in murine plasmacytomas and is rearranged similarly in human Burkitt lymphomas.
Authors: Adams J M , Gerondakis S , Webb E , Corcoran L M , Cory S
Published: May 2006

Molecular cloning has recently established that the 15;12 chromosome translocations in murine plasmacytomas fuse DNA from chromosome 15 to the immunoglobulin heavy (H) chain locus, usually within the switch recombination region near the alpha constant region gene. We show here that the incoming DNA bears the cellular gene (c-myc) homologous to the oncogene (v-myc) of avian retrovirus MC29. In human Burkitt lymphomas bearing an 8;14 translocation, c-myc was also rearranged, apparently (in at least two cases) to an H chain switch recombination region (mu or alpha), and both products of a reciprocal chromosome exchange were detectable. Both the murine and human c-myc genes contain two exons homologous to v-myc, and additional 5' and 3' murine genomic segments (apparent exons) were defined by hybridization to c-myc mRNAs. In plasmacytomas, chromosome breakpoints fall near or within the 5' exon and apparently disrupt the normal c-myc transcriptional unit, because plasmacytoma c-myc mRNAs differ from the mRNA in lines without c-myc rearrangement. The translocated gene presumably has lost its normal 5' regulatory sequences and may well encode an altered myc polypeptide. We propose that altered expression of the c-myc gene, induced by translocation to an immunoglobulin locus, is a critical oncogenic event for these B lymphoid tumors. Two events may be required, because the plasmacytoma oncogene capable of transforming fibroblasts is not c-myc.
Complete mRNA coding sequence of the acetylcholine binding alpha-subunit of Torpedo marmorata acetylcholine receptor: a model for the transmembrane organization of the polypeptide chain.
Authors: Devillers-Thiery A , Giraudat J , Bentaboulet M , Changeux J P
Published: May 2006

A 1,350-base-pair-long cDNA clone, named p alpha-2, was isolated by hybridization to the previously characterized clone p alpha-1 and found to be specific for the alpha-subunit of the Torpedo marmorata acetylcholine receptor. The nucleotide sequences of both cDNA inserts were analyzed and the sequence of the complete coding region and part of the 5' and 3' untranslated regions of the alpha-chain mRNA was determined. The complete amino acid sequence of the alpha-chain precursor is presented and used to develop a model for the transmembrane organization of the polypeptide.
Evidence for the involvement of a cyclic AMP-independent protein kinase in the activation of soluble tyrosine hydroxylase from rat striatum.
Authors: Andrews D W , Langan T A , Weiner N
Published: May 2006

Activation of rat striatal tyrosine hydroxylase [TyrOHase; tyrosine monooxygenase; L-tyrosine, tetrahydropteridine:oxygen oxidoreductase (3-hydroxylating), EC 1.14.16.2] by ATP/Mg2+ and endogenous protein kinase can be produced without the addition of cAMP. This activation is not due to endogenous free catalytic subunit derived from cAMP-dependent protein kinase. In the presence of amounts of protein kinase inhibitor sufficient for complete inhibition of striatal cAMP-dependent protein kinase and the cAMP-mediated activation of TyrOHase, addition of ATP/Mg2+ results in an enhancement of TyrOHase activity. Enzyme activation does not occur when the nonhydrolyzable form of ATP, adenylyl imidodiphosphate, is substituted for ATP. When TyrOHase is assayed in the presence of ATP/Mg2+ and different concentrations of either tyrosine or 6-methyltetrahydropterin co-factor, a 2-fold increase in enzyme Vmax is demonstrable, with no change in the Km for either substrate or cofactor. In contrast, in the presence of cAMP and ATP/Mg2+, both an increase in Vmax and an enhanced affinity for pterin cofactor are demonstrable. In the latter circumstance, the 2-fold increase in Vmax can be attributed entirely to the action of cAMP-independent protein kinase. The addition of either EGTA or CaCl2 does not modify the effect seen in the presence of ATP, suggesting that the effect of ATP/Mg2+ is not mediated by a Ca2+-dependent protein kinase. These data support the existence of a cAMP-independent striatal protein kinase that can catalyze the activation of TyrOHase.
Nucleotide sequence of the human parathyroid hormone gene.
Authors: Vasicek T J , McDevitt B E , Freeman M W , Fennick B J , Hendy G N , Potts J T , Rich A , Kronenberg H M
Published: May 2006

From a lambda phage gene library we have isolated phage containing the gene encoding human preproparathyroid hormone. The phage were isolated by using both the plaque-hybridization technique and the in vivo recombination-selection technique. The human preproparathyroid hormone gene contains two intervening sequences that separate the gene into a 5' noncoding domain, a "prepro" sequence domain, and a domain containing the parathyroid hormone sequence and the 3' noncoding region. The gene is approximately 4,200 base pairs long. Restriction endonuclease analysis of human leukocyte DNA shows that the haploid human genome contains one copy of the preproparathyroid hormone gene. A 14-base-pair sequence of alternating purines and pyrimidines that has the potential of adopting the Z-DNA conformation lies 134 base pairs upstream from the presumed site of initiation of transcription.
Tyrosine-specific protein kinase activity is associated with the purified insulin receptor.
Authors: Kasuga M , Fujita-Yamaguchi Y , Blithe D L , Kahn C R
Published: May 2006

Highly purified human placental insulin receptors were obtained by sequential affinity chromatography on wheat germ agglutinin and insulin-agarose. The preparation had an insulin binding capacity of 4,700 pmol/mg of protein approaching theoretical purity. The purified receptor revealed three major bands of Mr 135,000, 95,000, and 52,000 in NaDodSO4/polyacrylamide gel electrophoresis after reduction by dithiothreitol. All three bands were immunoprecipitated by anti-insulin-receptor antibodies. When this preparation was incubated with [gamma-32P]ATP in the presence of MnCl2 (2 mM) and analyzed in NaDodSO4/acrylamide gel electrophoresis, only the Mr 95,000 band was labeled. Preincubation with several concentrations of insulin increased the 32P incorporation into this peptide in dose-dependent fashion, whereas insulin-like growth factors were approximately equal to 2% as potent and epidermal growth factor had little or no effect, consistent with their known affinities for the insulin receptor. Insulin stimulation of phosphorylation of the Mr 95,000 subunit of the receptor was observed also in immunoprecipitates of this highly purified insulin receptor by anti-insulin-receptor antibodies. Phosphoamino acid determination revealed only phosphotyrosine in both the basal and insulin-stimulated states. These data suggest that a tyrosine-specific protein kinase activity is closely associated with insulin receptor, and this may be important in the signal transmission required for insulin action.
Evidence for a multiple random coupling mechanism in the alpha-ketoglutarate dehydrogenase multienzyme complex of Escherichia coli: a computer model analysis.
Authors: Hackert M L , Oliver R M , Reed L J
Published: May 2006

A computer modeling system was used to analyze experimental data for inactivation of the Escherichia coli alpha-ketoglutarate dehydrogenase complex accompanying release of lipoic acid residues by lipoamidase and by trypsin [Stepp, L. R., Bleile, D. M., McRorie, D. K., Pettit, F. H. & Reed, L. J. (1981) Biochemistry 20, 4555-4560]. The results provide insight into the active-site coupling mechanism in the alpha-ketoglutarate dehydrogenase complex. The model studies indicate that the overall activity of the alpha-ketoglutarate dehydrogenase complex is influenced by redundancies and random processes that we describe as a multiple random coupling mechanism. More than one lipoyl moiety services each E1 subunit (alpha-ketoglutarate dehydrogenase, EC 1.2.4.2), and an extensive lipoyl-lipoyl interaction network for exchange of electrons and possibly acyl groups must also be present. The best fit between computed and experimental data was obtained with a model that has two lipoyl moieties servicing each E1 subunit and a lipoyl-lipoyl interaction network that links all lipoyl moieties on the E2 cube (dihydrolipoamide succinyltransferase, EC 2.3.1.61). The single lipoyl moiety on an E2 subunit is assumed to service the coenzyme A-dependent succinyltransferase site of that E2 subunit as well as an E3 subunit (dihydrolipoamide dehydrogenase, EC 1.6.4.3) if the latter is bound to that particular E2 subunit.
Activation of Chi, a recombinator, by the action of an endonuclease at a distant site.
Authors: Stahl M M , Kobayashi I , Stahl F W , Huntington S K
Published: May 2006

Chi is an Escherichia coli recombinator specific to the recBC pathway. In phage lambda, its activity is dependent on its orientation with respect to the orientation of the chromosome packaging origin, cos [Kobayashi, I., Murialdo, H., Crasemann, J.M., Stahl, M. M. & Stahl, F. W. (1982) Proc. Natl. Acad. Sci. USA 79, 5981-5985]. Chi in an inactive state is activated by the in vivo action of a restriction endonuclease at a distant site. This result supports a model for Chi activation in which a recombination machine enters a DNA duplex at an end and travels until it encounters a Chi properly oriented with respect to the direction of enzyme movement. Events are then initiated that lead to exchange in the neighborhood of Chi.
Cultured hepatocytes from mouse strains expressing high and low levels of the fourth component of complement differ in rate of synthesis of the protein.
Authors: Rosa P A , Shreffler D C
Published: May 2006

Primary hepatocyte cultures prepared from the livers of inbred strains of mice show different rates of synthesis for the fourth component of complement (C4). The relative amounts of C4 synthesized by hepatocyte cultures correspond to the relative plasma levels of C4 among strains, a trait determined by the S region of the H-2 major histocompatibility complex. There is no evidence for increased intracellular catabolism of C4 in hepatocyte cultures from a strain with a low level of plasma C4. Processing and secretion of C4 proceed very slowly in vitro relative to other hepatocyte proteins.
Chemically synthesized peptides of hepatitis B surface antigen duplicate the d/y specificities and induce subtype-specific antibodies in chimpanzees.
Authors: Gerin J L , Alexander H , Shih J W , Purcell R H , Dapolito G , Engle R , Green N , Sutcliffe J G , Shinnick T M , Lerner R A
Published: May 2006

Synthetic peptides, predicted from the nucleotide sequence of the S gene of hepatitis B virus were analyzed in terms of the established specificities of the hepatitis B surface antigen. The analysis indicated that the group-specific alpha antigen is composed of at least three nonoverlapping sequences and that a relatively hydrophilic region of the surface antigen protein, spanning amino acid residues 110-137, specifies the major d and y subtype system. The d/y subtype appears to depend on changes in one or more variable amino acids at positions 127, 131, and 134 of the hepatitis B surface antigen protein. Peptide 49 (consisting of amino acid sequences of the y subtype for the region 110-137), coupled to a carrier protein and mixed with an adjuvant, stimulated a brisk anti-y response in chimpanzees, the relevant model of human response to hepatitis B virus immunization and infection. Experimental challenge with homologous hepatitis B virus resulted in a pattern of partial protection. The results offer promise for the application of chemically synthesized peptides as vaccines in the prophylaxis of hepatitis B virus disease.
DNase I-hypersensitive sites in the chromatin of immunoglobulin kappa light chain genes.
Authors: Chung S Y , Folsom V , Wooley J
Published: May 2006

We have used DNase I as a probe to examine the chromatin structure of mouse immunoglobulin kappa light chain genes in rearranged and unrearranged chromosomes--i.e., in nuclei from myeloma cells and from brain and liver cells. Tissue-specific DNase I-hypersensitive sites are observed 0.7 and 1.7 kilobases upstream from the 5' end of the C kappa gene in the J kappa -C kappa intron region in myeloma nuclei but not in naked DNA or in brain or liver nuclei. In myeloma cells expressing one functional kappa light chain polypeptide, but with more than one rearranged allele, one DNase I-hypersensitive site is found 0.3 kilobases upstream from the start of the coding region of the V kappa sequence in both functionally rearranged and nonfunctionally rearranged alleles but not in cross-hybridizing V kappa genes in the germ line context. Thus, during the development of B lymphocytes, the commitment of immunoglobulin kappa light chain gene expression seems to be associated with the presence of DNase I hypersensitivities that reflect changes of chromosomal structure surrounding the single copy C kappa gene. In contrast, the germ line V kappa multigene family seems to be located in a chromosomal region that does not exhibit change of DNase I hypersensitivity in response to commitment of immunoglobulin expression; a V kappa gene acquires DNase I hypersensitivity only after it is translocated adjacent to the J kappa -C kappa intron region. The DNase I-hypersensitive site 5' to the V kappa sequence is similar in location to hypersensitive sites found for other eukaryotic genes and is probably associated with the promoter region. However, the DNase I-hypersensitive sites in the J kappa -C kappa intron region are not associated with any known promoters. In addition, the DNA sequences surrounding the C kappa-proximal DNase I-hypersensitive site have several stretches of homology with sequences within the 72-base-pair tandem repeat of simian virus 40, which has been shown to modulate the transcriptional activity of neighboring genes. This DNase I-hypersensitive site in the intron region may be significant for the differential expression of the translocated V kappa genes.
Folding autonomy of the kringle 4 fragment of human plasminogen.
Authors: Trexler M , Patthy L
Published: May 2006

Kringle 4, an 88-residue plasminogen fragment carrying a lysine-binding site, loses its affinity for lysine-Sepharose upon reductive cleavage of its disulfide bridges. Aerobic incubation of the reduced, denatured fragment results in the rapid restoration of the disulfide bonds with concomitant recovery of lysine-Sepharose affinity. The ability of the unfolded fragment to regain its native conformation suggests that the kringle structure is an autonomous folding domain. During refolding of kringle 4 the native disulfide bonds, (formula; see text) and (formula; see text), appears first. The folding intermediate possessing these two disulfide bridges already binds to lysine-Sepharose, indicating that the third native bridge, which in native kringle 4 connects residues Cys1 and Cys79, is not essential for the maintenance of the biologically active conformation of kringle 4. Comparison of the sequences of human prothrombin, urokinase, and plasminogen kringles revealed that the residues surrounding the (formula; see text) and (formula; see text) bridges constitute the most conservative segments of kringles, whereas the residues neighboring the (formula; see text) bridge are not highly conserved. We propose that conservation of various residues in the different kringles reflects their importance for the folding autonomy of kringles.
Nucleotide sequence analysis of the proviral genome of avian myelocytomatosis virus (MC29).
Authors: Reddy E P , Reynolds R K , Watson D K , Schultz R A , Lautenberger J , Papas T S
Published: May 2006

The nucleotide sequence of the integrated proviral genome of avian myelocytomatosis virus (MC29) coding for gag-myc protein has been determined. By comparison of this nucleotide sequence with the helper virus as well as the c-myc region, it was possible to localize the junction points between helper viral and v-myc sequences. These studies demonstrate that (i) the large terminal repeat sequence of MC29 is very similar to that of Rous sarcoma virus, (ii) the viral genome has suffered extensive deletions in the gag, pol, and env genes, (iii) the gag region can code for p19, p10, and part of p27, (iv) the recombination between viral and cellular sequences occurred in the coding region of p27 such that the open reading frame extends for an additional stretch of 1,266 base pairs, resulting in a gag-myc hybrid protein, (v) the open reading frame terminated within the v-myc region 300 bases upstream of v-myc-helper viral junction, and (vi) the v-myc helper-viral junction at the 3' end occurred in the middle of env gene, rendering it defective.