bluuebunny/crossref_metadata_2025 huggingface dataset:
Why do polyethylene crystals have sectors?
Authors: Dorset D L , Hanlon J , McConnell C H , Fryer J R , Lotz B , Wittmann J C , Beckmann E , Zemlin F
Published: May 2006

High-resolution (3.7 A in optical diffraction) electron microscope images have been obtained from a series of n-paraffin monolamellar crystals with chain lengths from n-C36H74 to n-C82H166. The higher molecular weight specimens, which do not undergo chain folding, form sectorized crystals and the molecular packing is found to include alternate bands of untilted and tilted chains along <130>. Their widths are consistent with those of Bragg fringe widths in bright-field images obtained at lower magnification. The chain tilt axis is near d*110. Lower molecular weight paraffins form nonsectorized crystals where the chains are generally untilted with occasional small inclinations around nonspecific axes. Surface decoration of the longer alkanes with polyethylene crystallites, first of all, reveals three preferred polyethylene crystal rod orientations ([100] plus two perpendicular to [110]) instead of the two commonly found for the lower alkane. Control studies on solid-solution crystals reveal that the third [100] orientation is a result of slight surface roughness due to unequal chain lengths or surface protrusions of chains; the new decoration is also randomly distributed. For pure n-C60H122 lamellae, however, suggestions of regular bands containing rods along [100], due to surface discontinuities along <130>, can also be seen. In contrast with polyethylene, these data suggest that crystal sectorization may be a function of chain-stem packing alone and that chain folds may play merely a secondary role in the polymer--e.g., by directing the collapse of pyramidal crystals on a flat surface.
Promoter switching in Epstein-Barr virus during the initial stages of infection of B lymphocytes.
Authors: Woisetschlaeger M , Yandava C N , Furmanski L A , Strominger J L , Speck S H
Published: May 2006

Transcription of the genes encoding the six viral nuclear antigens present in Epstein-Barr virus latently infected lymphocytes can be initiated from one of two promoters (Cp and Wp) mapping near the left end of the viral genome. These promoters are used in a mutually exclusive manner in clonal cell lines established from either Burkitt lymphoma tumors or in vitro infection of peripheral B lymphocytes. In this paper the role of Cp and Wp during viral latency is investigated. Cp appears to be the promoter normally employed during established latent infection. Analysis of two cell lines that use Wp revealed a deletion spanning Cp in the endogenous viral genomes, suggesting that cell lines exhibiting Wp activity harbor mutated viral genomes with a nonfunctional Cp. However, in contrast to the preferred usage of Cp exhibited by established Epstein-Barr virus-infected cell lines, Wp was shown to be exclusively utilized during the initial stages of viral infection. In addition to Wp activity, Cp usage was apparent by 6 days post-infection. A model is proposed involving B-lymphocyte differentiation-driven promoter switching during the establishment of viral latency.
cDNA sequence for human erythrocyte ankyrin.
Authors: Lambert S , Yu H , Prchal J T , Lawler J , Ruff P , Speicher D , Cheung M C , Kan Y W , Palek J
Published: May 2006

The cDNA for human erythrocyte ankyrin has been isolated from a series of overlapping clones obtained from a reticulocyte cDNA library. The composite cDNA sequence has a large open reading frame of 5636 base pairs (bp) with the complete coding sequence for a polypeptide of 1879 amino acids with a predicted molecular mass of 206 kDa. The derived amino acid sequence contained 194 residues that were identical to those obtained by direct amino acid sequencing of 11 ankyrin proteolytic peptides. The primary sequence contained 23 highly homologous repeat units of 33 amino acids within the 90-kDa band 3 binding domain. Two cDNA clones showed evidence of apparent mRNA processing, resulting in the deletions of 486 bp and 135 bp, respectively. The 486-bp deletion resulted in the removal of a 16-kDa highly acidic peptide, and the smaller deletion had the effect of altering the COOH terminus of the molecule. Radiolabeled ankyrin cDNAs recognized two erythroid message sizes by RNA blot analysis, one of which was predominantly associated with early erythroid cell types. An ankyrin message was also observed in RNA from the human cerebellum by the same method. The ankyrin gene is assigned to chromosome 8 using genomic DNA from a panel of sorted human chromosomes.
Regulation of mRNA abundance in activated T lymphocytes: identification of mRNA species affected by the inhibition of protein synthesis.
Authors: Coleclough C , Kuhn L , Lefkovits I
Published: May 2006

Inhibition of protein synthesis has often been observed to increase the concentration of mRNAs that encode proteins associated with the regulation of cell division. As two-dimensional gel electrophoresis permits the simultaneous monitoring of individual elements in large populations of gene products, we have used this technique to assess the effect of cycloheximide treatment on the mRNA complement of activated mouse T cells in an objective fashion. Two-dimensional gels of proteins generated by cell-free translation of mRNA from T-cell blasts display about 400 spots; only 5 of these are reproducibly enhanced by cycloheximide treatment and about 4 are diminished. The cDNA cloning vector lambda jac allows analysis of large arrays of molecular clones by cell-free expression, and we have used it in a sibling selection scheme to isolate a clone of one of the prominently induced mRNA species, which we refer to as chx1. chx1 mRNA concentration is increased by cycloheximide treatment of activated B cells, as well as T cells, and it is rapidly and transiently induced, in a cycloheximide-enhanced manner, upon serum stimulation of resting 3T3 fibroblastoid cells. The chx1 protein is hydrophilic, is slightly basic, and has patches of homology with the Jun-D gene product. The chx1 gene is remarkable in its lack of detectable introns and of strong bias against CpG dinucleotides.
Transgenic mice expressing a soluble foreign H-2 class I antigen are tolerant to allogeneic fragments presented by self class I but not to the whole membrane-bound alloantigen.
Authors: Arnold B , Messerle M , Jatsch L , Küblbeck G , Koszinowski U
Published: May 2006

The properties of transmembrane and soluble transplantation antigens were compared with respect to the induction of tolerance and the selection of the T-cell repertoire. For this purpose, transgenic (H-2b x H-2d)F1 mice were constructed that carry integrated copies of a modified H-2Kk gene resulting in the secretion from various cell types including thymocytes of soluble H-2Kk molecules. Despite the presence of H-2Kk antigen, these mice were still able to generate an H-2Kk-specific T-cell response. This response was comparable to that produced by normal littermates when stimulated with cells expressing membrane H-2Kk in a mixed lymphocyte reaction. In contrast, only transgenic mice failed to generate a cytolytic T-cell response to soluble H-2Kk antigen expressed by recombinant vaccinia virus and presented by the H-2Db molecule. These data imply the presence of two populations of alloreactive cytolytic T cells. A small fraction of T cells recognizes alloantigen as antigenic peptide(s) presented by other major histocompatibility complex class I molecules and tolerance can be induced in this population by soluble alloantigen. The majority of T cells, however, require the whole cell membrane-expressed class I molecule for recognition. This population is not affected by tolerance induction to the soluble major histocompatibility complex class I molecule.
Self heat shock and gamma delta T-cell reactivity.
Authors: Rajasekar R , Sim G K , Augustin A
Published: May 2006

We have investigated the effects of heat shock on T-cell induction and selection in vitro. We find that when cell preparations containing T lymphocytes are incubated for 30 min at 42 degrees C, a selective proliferation of gamma delta + T cells bearing the gamma delta T-cell antigen receptor follows. A greater enrichment of gamma delta + T cells is observed, upon preexposure to mycobacterial antigens in vivo. By comparing the effects of heat shock with that of mitogen or specific T-cell triggering by conventional antigens and by analyzing the gamma delta T-cell receptor genes expressed in cells that proliferate as a result of heat shock induction, we conclude that a subset of murine gamma delta T cells react to antigens on self cells in which a heat shock response was induced.
Cytoskeletal reorganizations responsible for the phorbol ester-induced formation of cytoplasmic processes: possible involvement of intermediate filaments.
Authors: Bershadsky A D , Ivanova O Y , Lyass L A , Pletyushkina O Y , Vasiliev J M , Gelfand I M
Published: May 2006

The tumor promoter phorbol 12-myristate 13-acetate (PMA) induces characteristic reversible changes of cell shape in certain fibroblastic lines: motile lamellas are transformed into noncontractile narrow processes; simultaneously, the actin microfilament network of lamellas is locally disorganized. This reaction to PMA may be regarded as a prototype of reorganizations involving formation of stable cytoplasmic processes. Specific drugs, Taxol and Colcemid, were used to study the role of microtubules and vimentin-containing intermediate filaments (IF) in the development of PMA-induced reorganizations. PMA readily induced formation of noncontractile processes in Taxol-treated fibroblasts; these cells had a profoundly altered microtubular system but noncollapsed IF. A short (1 hr) exposure to PMA induced formation of processes in control cells but not in the Colcemid-treated cells, which had depolymerized microtubules and IF that collapsed around the nucleus. Longer (3-4 hr) exposure of the Colcemid-treated cells to PMA induced partial reversal of the IF collapse; those parts of the peripheral lamellas that contained IF were transformed into narrow noncontractile processes. It is suggested that the local interaction of IF with the actin system is an essential step in the formation of processes from lamellas. The microtubular system controls distribution of IF in the cytoplasm and thus plays an indirect role in the reorganization of the actin cortex.
Identification of an enzyme in human kidney that correctly processes prorenin.
Authors: Shinagawa T , Do Y S , Baxter J D , Carilli C , Schilling J , Hsueh W A
Published: May 2006

Using pure recombinant human prorenin as a substrate, we have identified an enzyme in human kidney that accurately processes prorenin to active renin (EC 3.4.23.15). In the crude homogenate, the predominant activity of this potential renin-processing enzyme (RPE) converted the Mr 47,000 inactive prorenin to Mr 44,000 active renin and had a pH optimum of approximately 6. The activity was blocked by cysteine protease inhibitors, but not by pepstatin, EDTA, or serine protease inhibitors. This RPE activity was not detected in a similarly prepared homogenate of human chorion decidua tissue, which produces primarily prorenin, or in human plasma. The activity was purified 100-fold by ammonium sulfate precipitation, p-chloromercuribenzoate affinity chromatography, and chromatofocusing. The partially purified enzyme has a Mr of approximately 27,000 and an isoelectric point in the pH 4.8-5.6 range. The activity in the purified RPE preparation had the same pH optimum as that in crude homogenate, cleaved the prosegment at the same site used by the kidney in vivo based on amino-terminal sequencing of the processed renin, and did not degrade prorenin or renin. These data suggest that the cysteine protease we have isolated is a candidate for authentic renal RPE.
Habituation in the single cell: diminished secretion of norepinephrine with repetitive depolarization of PC12 cells.
Authors: McFadden P N , Koshland D E
Published: May 2006

Neuronally differentiated PC12 cells secrete decreasing amounts of [3H]norepinephrine when repetitively stimulated by depolarizing concentrations of potassium ion. The decreasing response shows attributes that have been classically ascribed to response habituation, a behavior commonly observed in nervous systems but found here in a homogeneous cell type. Alteration of the habituation pattern was caused by activators of the protein kinase C pathway and of voltage-gated calcium channels.
Hepatitis C virus shares amino acid sequence similarity with pestiviruses and flaviviruses as well as members of two plant virus supergroups.
Authors: Miller R H , Purcell R H
Published: May 2006

Hepatitis C virus (HCV) is an important human pathogen that is associated with transfusion-related non-A, non-B hepatitis. Recently, HCV cDNA was cloned and the nucleotide sequence of approximately three-quarters of the virus genome was determined. A region of the predicted polyprotein sequence was found to share similarity with a nonstructural protein encoded by dengue virus, a member of the flavivirus family. We report here that HCV shares an even greater degree of protein sequence similarity with members of the pestivirus group (i.e., bovine viral diarrhea virus and hog cholera virus), which are thought to be distantly related to the flaviviruses. In addition, we find that HCV shares significant protein sequence similarity with the polyproteins encoded by members of the picornavirus-like and alphavirus-like plant virus supergroups. These data suggest that HCV may be evolutionarily related to both plant and animal viruses.
Recognition by CD8 on cytotoxic T lymphocytes is ablated by several substitutions in the class I alpha 3 domain: CD8 and the T-cell receptor recognize the same class I molecule.
Authors: Connolly J M , Hansen T H , Ingold A L , Potter T A
Published: May 2006

The CD8 molecule on class I-reactive cytotoxic T lymphocytes (CTLs) is believed to function as a coreceptor along with the alpha beta T-cell receptor. Whereas the alpha beta T-cell receptor recognizes polymorphic residues in the alpha 1/alpha 2 domains of the class I molecule, the CD8 molecule is believed to recognize monomorphic class I residues. Our previous experiments suggested that residue 227 in the alpha 3 domain of major histocompatibility complex class I molecules contributes to the determinant recognized by CD8. By using a panel of site-directed mutants of H-2Dd, this observation has been extended herein. Our findings indicate that for recognition by CD8-dependent CTLs, residue 227 must be either glutamic acid or aspartic acid and cannot be either basic or uncharged. However, the recognition by CD8-independent CTLs is unaffected by any of the substitutions at position 227 of H-2Dd. Similarly, alterations of other charged residues at positions 222, 223, and 229 have an analogous effect to substitution at residue 227, whereas substitutions at residues 192 and 232 do not affect the reactivity of CD8-dependent or CD8-independent CTLs. In addition, mutant H-2Dd molecules that are not recognized by CD8-dependent CTLs are unable to stimulate a primary CTL response, yet they can stimulate a secondary CD8-independent H-2Dd-specific CTL response. These findings suggest that CD8 recognition is obligatory for the priming of class I-dependent CTL responses. Since endogenous class I molecules were expressed by all of the transfected cell lines, these findings provide direct genetic evidence that CD8 and the alpha beta T-cell receptor must interact with the same class I molecule.
Neuropeptide stimulation of calcium flux in human lung cancer cells: delineation of alternative pathways.
Authors: Bunn P A , Dienhart D G , Chan D , Puck T T , Tagawa M , Jewett P B , Braunschweiger E
Published: May 2006

Calcium ion flux following the administration of a series of neuropeptides, N6,O2'-dibutyryladenosine 3',5'-cyclic monophosphate, and serum was monitored by flow cytometry in selected lung and breast cancer cell lines and Chinese hamster ovary cell line CHO-K1. Calcium ion flux was monitored in individual cells by flow cytometry using the indicator indo-1 AM. Five groups of neuropeptides produced calcium flux changes in lung cancer cell lines and CHO-K1 cells but not in breast cancer cells. The peak increase in free calcium was reached within 10 sec of peptide administration and declined to resting levels in 70-120 sec. When two or more members of the same group were administered simultaneously, calcium flux changes were identical to that produced by each single peptide. When two or more members of different groups were administered simultaneously, an increased calcium release occurred. When identical peptides or peptides from the same group were administered sequentially after the return of calcium concentrations to resting values, no calcium flux resulted from the second peptide. When peptides from different active groups were administered sequentially, a new calcium flux occurred after each peptide. These data are interpreted to mean that members of each active group of peptides trigger a different calcium flux pathway. Thus, many such pathways and different metabolic states exist within the cell. Elucidation of calcium flux pathways in normal and cancer cells may lead to greater understanding of the nature of the malignant defect.
The alpha 1 (IX) collagen gene gives rise to two different transcripts in both mouse embryonic and human fetal RNA.
Authors: Muragaki Y , Nishimura I , Henney A , Ninomiya Y , Olsen B R
Published: May 2006

We have isolated and characterized portions of the alpha 1 (IX) collagen gene from mouse and human DNA. Nucleotide sequence analysis and comparison with the chicken gene suggest that the mammalian genes contain an alternative exon that is located within the intron between exons 6 and 7. Using oligonucleotide primers specific for exons 4, 8, and the alternative exon (exon 1*), we demonstrated by the polymerase chain reaction that embryonic mouse and fetal human RNAs contain two types of alpha 1(IX) collagen transcripts. One type of transcript does not contain the sequence encoded by exon 1*; the second type of transcript contains this exon. Both mouse and human alpha 1(IX) collagen genes give rise, therefore, to (at least) two mRNA transcripts.
Presentation of human minor histocompatibility antigens by HLA-B35 and HLA-B38 molecules.
Authors: Yamamoto J , Kariyone A , Akiyama N , Kano K , Takiguchi M
Published: May 2006

Cytotoxic T lymphocyte (CTL) clones specific for human minor histocompatibility antigens (hmHAs) were produced from a patient who had been grafted with the kidneys from his mother and two HLA-identical sisters. Of eight CTL clones generated, four recognized an hmHA (hmHA-1) expressed on cells from the mother and sister 3 (second donor); two recognized another antigen (hmHA-2) on cells from the father, sister 2 (third donor), and sister 3; and the remaining two clones recognized still another antigen (hmHA-3) on cells from the father and sister 3. Panel studies revealed that CTL recognition of hmHA-1 was restricted by HLA-B35 and that of hmHA-2 and hmHA-3 was restricted by HLA-B38. The HLA-B35 restriction of the hmHA-1-specific CTL clones was substantiated by the fact that they killed HLA-A null/HLA-B null Hmy2CIR targets transfected with HLA-B35 but not HLA-B51, -Bw52, or -Bw53 transfected Hmy2CIR targets. These data demonstrated that the five amino acids substitutions on the alpha 1 domain between HLA-B35 and -Bw53, which are associated with Bw4/Bw6 epitopes, play a critical role in the relationship of hmHA-1 to HLA-B35 molecules. The fact that the hmHA-1-specific CTLs failed to kill Hmy2CIR cells expressing HLA-B35/51 chimeric molecules composed of the alpha 1 domain of HLA-B35 and other domains of HLA-B51 indicated that eight residues on the alpha 2 domain also affect the interaction of hmHA-1 and the HLA-B35 molecules.
Inhibition of leukocyte adherence by anti-CD18 monoclonal antibody attenuates reperfusion injury in the rabbit ear.
Authors: Vedder N B , Winn R K , Rice C L , Chi E Y , Arfors K E , Harlan J M
Published: May 2006

Tissue injury resulting from ischemia and reperfusion forms the basis of several important disorders including myocardial infarction, stroke, and circulatory shock. To examine the role of neutrophils in this process and to determine the extent to which injury is a consequence of reperfusion, we utilized the monoclonal antibody 60.3, directed to CD18, the human leukocyte adherence glycoprotein, to block intravascular neutrophil aggregation and neutrophil adherence to endothelium in a rabbit model of tissue ischemia and reperfusion. Antibody treatment either before ischemia or after ischemia, but prior to reperfusion, resulted in the same degree of significant protection against endothelial, microvascular, and tissue injury. We conclude that neutrophils and increased neutrophil adhesiveness are important in the development of microvascular and tissue injury after ischemia and reperfusion and that under these circumstances, injury is primarily a consequence of reperfusion.
Molecular cloning of a plant betaine-aldehyde dehydrogenase, an enzyme implicated in adaptation to salinity and drought.
Authors: Weretilnyk E A , Hanson A D
Published: May 2006

Many plants, as well as other organisms, accumulate betaine (N,N,N-trimethylglycine) as a nontoxic or protective osmolyte under saline or dry conditions. In plants, the last step in betaine synthesis is catalyzed by betaine-aldehyde dehydrogenase (BADH, EC 1.2.1.8), a nuclear-encoded chloroplastic enzyme. A cDNA clone for BADH (1812 base pairs) was selected from a lambda gt10 cDNA library derived from leaves of salt-stressed spinach (Spinacia oleracea L.). The library was screened with oligonucleotide probes corresponding to amino acid sequences of two peptides prepared from purified BADH. The authenticity of the clone was confirmed by nucleotide sequence analysis; this analysis demonstrated the presence of a 1491-base-pair open reading frame that contained sequences encoding 12 peptide fragments of BADH. The clone hybridized to a 1.9-kilobase mRNA from spinach leaves; this mRNA was more abundant in salt-stressed plants, consistent with the known salt induction of BADH activity. The amino acid sequence deduced from the BADH cDNA sequence showed substantial similarities to those for nonspecific aldehyde dehydrogenases (EC 1.2.1.3 and EC 1.2.1.5) from several sources, including absolute conservation of a decapeptide in the probable active site. Comparison of deduced and determined amino acid sequences indicated that the transit peptide may comprise only 7 or 8 residues, which is atypically short for precursors to stromal proteins.
Differentiation of thymocytes from CD3-CD4-CD8- through CD3-CD4-CD8+ into more mature stages induced by a thymic stromal cell clone.
Authors: Tatsumi Y , Kumanogoh A , Saitoh M , Mizushima Y , Kimura K , Suzuki S , Yagi H , Horiuchi A , Ogata M , Hamaoka T
Published: May 2006

We have investigated the capacity of our established thymic stromal cell clone (MRL104.8a) or its derived factor(s) to induce the differentiation of immature thymocytes. Culture of purified adult murine double-negative (CD4-CD8-, indicated here as CD4-8-) thymocytes on the MRL104.8a thymic stromal cell monolayer for 1 day resulted in the induction of an appreciable percentage of CD4-8+ thymocytes. A bone marrow-derived stromal cell monolayer or a L929 fibroblast monolayer failed to generate CD4-8+ cells. This differentiation could also be induced by a semipurified sample of the MRL104.8a culture supernatant, which contained a thymic stroma-derived T-cell growth factor capable of contributing to the growth of double-negative immature thymocytes. CD4-8+ thymocytes generated 1 day after coculture with the MRL104.8a cells or the sample containing thymic stroma-derived T-cell growth factor were found to be CD3- and J11d+, excluding the possibility of expansion of mature (CD3+4-8+) thymocytes present in the thymus. More importantly, when the culture period was extended to 2 or 3 days, an appreciable number of CD4+8+ and single-positive (CD4+) cells were generated on the MRL104.8a monolayer. Thus, these results provide the direct demonstration that CD3-4-8- immature thymocytes are promoted to differentiate through a rapidly cycling intermediate (CD3-4-8+) into double- and single-positive cells by a specialized thymic stromal component.
Epithelial glycoprotein is a member of a family of epithelial cell surface antigens homologous to nidogen, a matrix adhesion protein.
Authors: Simon B , Podolsky D K , Moldenhauer G , Isselbacher K J , Gattoni-Celli S , Brand S J
Published: May 2006

The cell surface antigen, epithelial glycoprotein, defined by the monoclonal antibody HEA 125, is expressed on virtually all epithelial cell membranes but not on mesodermal or neural cell membranes. The cDNA encoding epithelial glycoprotein was isolated by HEA 125 antibody enrichment of colon tumor cDNA expressed transiently in COS cells. The sequence of the epithelial glycoprotein antigen is identical to the cell membrane protein recognized by the monoclonal antibody KS 1/4 and is homologous to the tumor-associated antigen GA733. These proteins share sequence homology to nidogen, an extracellular matrix component that appears to participate in cell-matrix adhesion. These proteins also share a homologous domain found in the B1 chain of laminin, a matrix adhesion protein, and placental protein 12, an insulin-like growth factor I binding protein secreted during pregnancy that has been implicated in regulation of fetal growth. This common domain is also repeated multiple times within the thyroglobulin precursor. These findings suggest epithelial glycoprotein is a cell surface molecule involved in cell-cell or cell-matrix interaction.
cDNA cloning and expression of a metalloproteinase inhibitor related to tissue inhibitor of metalloproteinases.
Authors: Boone T C , Johnson M J , De Clerck Y A , Langley K E
Published: May 2006

The purification and characterization of a metalloproteinase inhibitor (MI) from bovine aortic endothelial cells, and the demonstration that it is related to, but distinct from, tissue inhibitor of metalloproteinase (TIMP), have previously been reported [De Clerck, Y. A., Yean, T.-D., Ratzkin, B. J., Lu, H.S. & Langley, K. E. (1989) J. Biol. Chem. 264, 17445-17453]. The cDNA cloning of the bovine MI and its human homolog is now reported. The bovine cDNA cloning used probes designed on the basis of NH2-terminal amino acid sequence of bovine MI. The human cDNA cloning in turn used probes representing parts of the bovine cDNA nucleotide sequence. Both cDNAs encode leader sequences of 26 amino acids and mature protein sequences of 194 amino acids. The amino acid sequences of the mature proteins are 94% identical. The human MI cDNA was expressed in Escherichia coli, and a preparation containing anticollagenase activity was recovered. The amino acid sequence of mature human MI is 38% identical to the sequence for human TIMP, and the 12 cysteines in MI and TIMP are aligned almost identically. Thus MI and TIMP comprise an inhibitor family.
Organ-specific transcripts of different size and abundance derive from the same pyruvate, orthophosphate dikinase gene in maize.
Authors: Glackin C A , Grula J W
Published: May 2006

Analyses of genomic DNA and clones indicate that the pyruvate, orthophosphate dikinase (PPDK; ATP: pyruvate, orthophosphate phosphotransferase, EC 2.7.9.1) gene family of maize (Zea mays L. subsp. mays, line B73) contains two members. Restriction site and DNA sequence comparisons between PPDK genomic and leaf cDNA clones have revealed which gene encodes the isozyme involved in C4 photosynthesis. The region flanking the 5' end of this gene contains two 30-base-pair (bp) repetitive elements that may be involved in its light-regulated expression. Sequence analysis of genomic and leaf cDNA clones has also shown that the entire 7.3-kDa PPDK chloroplast transit peptide is encoded in the 436-bp first exon. Northern blot experiments with probes specific for the first exon and the 3' end of the gene showed that the smaller PPDK transcripts in roots and etiolated leaves [3.0 kilobases (kb) vs. the 3.5-kb green leaf transcript] lack the sequence encoding the chloroplast transit peptide. In addition, results from cDNA library screens have confirmed that the root transcript is approximately 50-fold less abundant than the green leaf transcript. Finally, sequence comparisons among cDNA clones from green leaves and roots and genomic clones representing both members of the PPDK gene family demonstrate that the green leaf transcript encoding the C4 isozyme and the root transcript are derived from the same gene.
Suppression of feline immunodeficiency virus infection in vivo by 9-(2-phosphonomethoxyethyl)adenine.
Authors: Egberink H , Borst M , Niphuis H , Balzarini J , Neu H , Schellekens H , De Clercq E , Horzinek M , Koolen M
Published: May 2006

The acyclic purine nucleoside analogue 9-(2-phosphonomethoxyethyl)adenine [PMEA; formerly referred to as 9-(2-phosphonylmethoxyethyl)adenine] is a potent and selective inhibitor of human immunodeficiency virus replication in vitro and of Moloney murine sarcoma virus-induced tumor formation in mice. In the latter system PMEA has stronger antiretroviral potency and selectivity than 3'-azido-3'-thymidine (AZT). We have now investigated the effect of the drug in cats infected with the feline immunodeficiency virus (FIV). In vitro, PMEA was found to efficiently block FIV replication in feline thymocytes (50% effective dose, 0.6 microM). When administered to cats at doses of 20, 5, or 2 mg/kg per day, PMEA caused a dose-dependent suppression of FIV replication and virus-specific antibody production. Seropositive field cats with signs of opportunistic infection (gingivitis, stomatitis, and diarrhea) showed clinical improvement during PMEA therapy (5 mg/kg per day) and recurrence of the disease after treatment was discontinued. Thus, FIV infection in cats is an excellent model to test the efficacy of selective anti-human immunodeficiency virus agents in vivo.
Solubilization and functional reconstitution of the protein-translocation enzymes of Escherichia coli.
Authors: Driessen A J , Wickner W
Published: May 2006

The SecY protein and other membrane proteins of Escherichia coli were solubilized by mixed micelles of n-octyl beta-D-glucopyranoside, phospholipids, and glycerol. Proteoliposomes formed from this extract by detergent dialysis supported energy-dependent translocation and processing of pro-OmpA. Translocation required ATP, SecY, and SecA and was stimulated by a proton-motive force. These results provide an important assay for the isolation and identification of membrane components involved in protein translocation.
More cold fusion in metals: corrected calculations and other considerations.
Authors: Parmenter R H , Lamb W E
Published: May 2006

A simple model of a metal containing deuterons is considered. The example of palladium is treated in detail. In contrast to a previous calculation, it is shown that the effect of screening of Coulomb fields by conduction electrons is sufficient to allow a deuteron pair to fuse at a rate of 10(-18) sec-1, five powers of 10 larger than some experimental results. It is shown that the Oppenheimer-Phillips process enhances the rate by a factor of 1.77. Difficulties with applying the model to titanium are discussed.
Molybdopterin guanine dinucleotide: a modified form of molybdopterin identified in the molybdenum cofactor of dimethyl sulfoxide reductase from Rhodobacter sphaeroides forma specialis denitrificans.
Authors: Johnson J L , Bastian N R , Rajagopalan K V
Published: May 2006

The nature of molybdenum cofactor in the bacterial enzyme dimethyl sulfoxide reductase has been investigated by application of alkylation conditions that convert the molybdenum cofactor in chicken liver sulfite oxidase and milk xanthine oxidase to the stable, well-characterized derivative [di(carboxamidomethyl)]molybdopterin. The alkylated pterin obtained from dimethyl sulfoxide reductase was shown to be a modified form of alkylated molybdopterin with increased absorption in the 250-nm region of the spectrum and altered chromatographic behavior. The complex alkylated pterin was resolved into two components by treatment with nucleotide pyrophosphatase. These were identified as di(carboxamidomethyl)molybdopterin and GMP by their absorption spectra, coelution with standard compounds, and by further degradation by alkaline phosphatase to dephospho [di(carboxamidomethyl)]molybdopterin and guanosine. The GMP moiety was sensitive to periodate, identifying it as the 5' isomer. Chemical analysis of the intact alkylated pterin showed the presence of two phosphate residues per pterin. These results established that the pterin isolated from dimethyl sulfoxide reductase contains the phosphoric anhydride of molybdopterin and 5'-GMP, which is designated molybdopterin guanine dinucleotide.
Transposition rates of movable genetic elements in Drosophila melanogaster.
Authors: Harada K , Yukuhiro K , Mukai T
Published: May 2006

A considerable proportion of visible mutations is reported to be caused by the insertion of mobile genetic elements in Drosophila and other organisms. We estimated transposition rates of some Drosophila mobile elements by using the lines AW and JH in which spontaneous mutations have been accumulated independently for about 400 generations. Occupied sites of the mobile elements were detected by in situ hybridization on the salivary gland chromosomes sampled from 40 AW and 30 JH lines. The rates of insertion and excision of the copia and two copia-like elements, 412 and 17.6, are very low: Insertions occurred at up to 10(-3) per second chromosome per generation (17.6) and excision occurred at about 10(-5) per site per generation (copia and 412). Insertions of the I and hobo elements occurred much more frequently. These estimates are not only important for assessing the actual rate of various types of mutations but also for developing an evolutionary theory of mobile elements themselves.
Interleukin 6 dependence of spontaneous in vitro differentiation of B cells from patients with IgM gammapathy.
Authors: Levy Y , Fermand J P , Navarro S , Schmitt C , Vainchenker W , Seligmann M , Brouet J C
Published: May 2006

Blood B cells from eight patients with clonal lymphoid disorders characterized by monoclonal IgM secretion (four with malignant plasmacytic proliferation typical of Waldenström macroglobulinemia and four without overt lymphoid neoplasia) were found to spontaneously differentiate in vitro into plasma cells. In all instances, monoclonal plasma cells (8-45% of the cells) were generated from extensively purified B cells or T-cell-depleted peripheral blood mononuclear cells after a 7-day culture period, with a corresponding high rate of IgM secretion into the culture medium. This differentiation occurred in the absence of any cell proliferation process as measured by [3H]thymidine uptake at day 2 or 4. Normal B cells did not differentiate under the same experimental conditions. Detection of interleukin 6 (IL-6) bioactivity in all patients' B-cell culture supernatants as well as of IL-6 mRNA in freshly prepared, uncultured B cells in the two cases studied by in situ hybridization suggested that IL-6 secretion by B cells may play a role in this process. Moreover, in the four patients without overt lymphoid proliferation, B-cell differentiation was significantly inhibited (60-80%) in the presence of anti-IL-6 antibodies. In contrast, anti-IL-6 antibodies did not preclude the differentiation into plasma cells of B cells from the four patients with bona fide Waldenström macroglobulinemia. These results suggest a two-step pathogenesis for such human lymphoplasmacytic clonal proliferations, the initial stage being characterized by an IL-6-dependent autocrine differentiation pathway.
The insulin receptor with phenylalanine replacing tyrosine-1146 provides evidence for separate signals regulating cellular metabolism and growth.
Authors: Wilden P A , Backer J M , Kahn C R , Cahill D A , Schroeder G J , White M F
Published: May 2006

We have studied the function of a mutant insulin receptor (IR) molecule in which Tyr-1146, one of the first autophosphorylation sites in the beta subunit, was replaced with phenylalanine (IRF1146). Autophosphorylation of the partially purified IRF1146 was reduced 60-70% when compared to the wild-type IR but was still stimulated by insulin. The phosphotransferase activity of the dephospho form of both the IR and IRF1146 toward exogenous substrates was stimulated 3- to 4-fold by insulin. However, the wild-type IR was activated 12-fold by autophosphorylation, whereas the IRF1146 was activated only 2-fold. When the IRF1146 was expressed in Chinese hamster ovary (CHO) cells, insulin binding was normal, whereas autophosphorylation was reduced 80% when compared to cells expressing the wild-type IR. Endogenous substrates of the insulin receptor kinase were not detected during insulin stimulation of CHO cells expressing the IRF1146. Moreover, the IRF1146 did not internalize insulin rapidly or stimulate DNA synthesis in the presence of insulin. In contrast, both the IR and IRF1146 stimulated glycogen synthase equally in CHO cells. These data suggest that activation of the IR tyrosine kinase can be resolved into two components: the first is dependent on insulin binding and the second is dependent on the subsequent insulin-stimulated autophosphorylation cascade. Thus, at least two signal transduction pathways diverging from the IR are implicated in the mechanism of insulin action.
Lipoprotein lipaseBethesda: a single amino acid substitution (Ala-176----Thr) leads to abnormal heparin binding and loss of enzymic activity.
Authors: Beg O U , Meng M S , Skarlatos S I , Previato L , Brunzell J D , Brewer H B , Fojo S S
Published: May 2006

The molecular defect that leads to a deficiency of lipoprotein lipase (LPL) activity in the proband from a Bethesda kindred has been identified. The pre- and post-heparin plasma LPL mass in the proband was elevated when compared to controls; however, there was no detectable LPL activity, indicating the presence of a defective enzyme (termed LPLBethesda). Analysis of the patient's post-heparin plasma by heparin-Sepharose affinity chromatography demonstrated that the mutant LPL had an altered affinity for heparin. Southern blot hybridization of the gene for LPLBethesda revealed no major rearrangements. Northern blot analysis of LPLBethesda mRNA from patient monocyte-derived macrophages revealed normal-sized mRNAs (3.4 and 3.7 kilobases) as well as normal cellular mRNA levels when compared to control macrophages. Sequence analysis of polymerase chain reaction-amplified LPL cDNA revealed a G----A substitution at position 781 of the normal LPL gene that resulted in the substitution of an alanine for a threonine at residue 176 and the loss of an SfaNI site present in the normal LPL gene. Amplification of cDNA by the PCR followed by digestion with SfaNI established that the patient was a true homozygote for the mutation. Expression of LPL cDNA in COS-7 cells resulted in the synthesis of a nonfunctional LPL enzyme establishing that the Ala----Thr substitution was the mutation responsible for the inactive LPL. The identification of this mutation in the LPL gene defines a region of the LPL enzyme, at Ala-176, that is essential for normal heparin-binding and catalytic activity. We propose that an amino acid substitution in this critical region of LPLBethesda results in the synthesis of a nonfunctional enzyme that leads to the chylomicronemia syndrome expressed in this proband.
Autoantibodies from a patient with scleroderma CREST recognized kinetochores of the higher plant Haemanthus.
Authors: Mole-Bajer J , Bajer A S , Zinkowski R P , Balczon R D , Brinkley B R
Published: May 2006

Human autoantibodies from a patient with scleroderma CREST (calcinosis, Raynaud phenomenon, esophageal dismotility, sclerodactyly, telangiectasia) were used to immunostain kinetochores on chromosomes in endosperm of the seed of the monocot Haemanthus katherinae Bak. Kinetochores of mitotic chromosomes and prekinetochores of interphase cells were specifically stained using conventional indirect immunofluorescence procedures as well as a nonfading immunogold-silver-enhanced technique and analyzed by fluorescence and video microscopy. In interphase, prekinetochores were either single or double structures depending on the stage of the cell cycle but became quadruple (two distinct stained dots on each chromatid) in mid-to-late prophase. In favorable preparations of prometaphase chromosomes, multiple subunits could be resolved within each sister kinetochore suggesting a compound organization. Western blot analysis demonstrated common epitopes in centromeric peptides of HeLa and Haemanthus cell extracts. Although the molecular mass of individual polypeptides differed in the two species, the presence of shared epitopes indicates striking conservation of centromere/kinetochore components throughout evolution.
Drosophila retrotransposon promoter includes an essential sequence at the initiation site and requires a downstream sequence for full activity.
Authors: Jarrell K A , Meselson M
Published: May 2006

We describe a 98-base-pair region (-38 to +60) in the long terminal repeat of the Drosophila gypsy retrotransposon that is sufficient for accurate normal-level transcription. We find that, unlike most RNA polymerase II (pol II) promoters, the gypsy promoter includes downstream sequences that are required for full activity. Also unlike most pol II promoters, the gypsy promoter, which lacks a TATA motif, was found to have an essential sequence at the transcription initiation site, mutation of which abolishes transcription. These three uncommon features of the gypsy promoter may be characteristic of a subset of pol II promoters, exemplified by certain retrotransposons and developmental genes of Drosophila and by Tdt, the mouse terminal deoxynucleotidyl-transferase (TdT) gene.
Structural and functional conservation of histidinol dehydrogenase between plants and microbes.
Authors: Nagai A , Ward E , Beck J , Tada S , Chang J Y , Scheidegger A , Ryals J
Published: May 2006

The partial amino acid sequence of histidinol dehydrogenase (L-histidinol:NAD+ oxidoreductase, EC 1.1.1.23) from cabbage was determined from peptide fragments of the purified protein. The relative positions of these peptides were deduced by aligning their sequences with the sequence of the HIS4C gene product of Saccharomyces cerevisiae. cDNA encoding histidinol dehydrogenase was then amplified from a library using a polymerase chain reaction primed with degenerate oligonucleotide pools of known position and orientation. By using this amplified fragment as a probe, an apparently full-length cDNA clone was isolated that is predicted to encode a proenzyme having a putative 31-amino acid chloroplast transit peptide and a mature molecular mass of 47.5 kDa. The predicted protein sequence was 51% identical to the yeast enzyme and 49% identical to the Escherichia coli enzyme. Expression of the cDNA clone in an E. coli his operon deletion strain rendered the mutant able to grow in the presence of histidinol.
CpG island in the region of an autosomal dominant polycystic kidney disease locus defines the 5' end of a gene encoding a putative proton channel.
Authors: Gillespie G A , Somlo S , Germino G G , Weinstat-Saslow D , Reeders S T
Published: May 2006

In an attempt to isolate candidate genes for autosomal dominant polycystic kidney disease, a number of CpG-rich islands have been identified from a region defined genetically as the site of disease mutations. Genomic fragments adjacent to one of these islands were used to isolate cDNAs from both HeLa cells and cultured cystic epithelium that encode a 155-amino acid peptide having four putative transmembrane domains. The corresponding transcript was found in all tissues tested but was most abundant in brain and kidney. Potential control response elements were identified in the genomic region 5' to the initiation codon. The deduced amino acid sequence has 93% similarity to the 16-kDa proteolipid component that is believed to be part of the proton channel of the vacuolar H(+)-ATPase. Possible roles for a mutated proton channel in the pathogenesis of cystic disease were considered. However, sequencing of cDNAs corresponding to both alleles of an affected individual revealed no differences in the deduced amino acid sequence. Moreover, transcript size and abundance were not altered in cystic kidney.
Phosphatidylinositol-stimulated phosphorylation of an inhibitory subunit of cGMP phosphodiesterase in vertebrate rod photoreceptors.
Authors: Hayashi F , Lin G Y , Matsumoto H , Yamazaki A
Published: May 2006

An inhibitory subunit (P gamma) of cGMP phosphodiesterase from vertebrate rod photoreceptors (frog, toad, and bovine) was phosphorylated by cytosolic protein kinase(s) derived from intact frog rod outer segments. The phosphorylation of frog P gamma was stimulated by phosphatidylinositol but not by cAMP or cGMP. One- and two-dimensional gel electrophoresis revealed that 70-80% of P gamma was phosphorylated with 1 mol of phosphate per frog P gamma under optimal conditions. A peptide that derived from an active domain of bovine P gamma was also phosphorylated. Phosphorylation of frog P gamma was inhibited by addition of the peptide to the reaction mixture. Phosphorylation of frog P gamma was also inhibited by addition of transducin subunits or active (P gamma-less) cGMP phosphodiesterase. Okadaic acid, on the other hand, enhanced P gamma phosphorylation, suggesting the presence of protein phosphatase(s) in the cytosolic fraction. These data suggest another mechanism for the regulation of cGMP phosphodiesterase in vertebrate rod photoreceptors.
Analysis of the energy of the first four excited states of the ground- state rotational bands of the even-even lanthanon nuclei (58Ce to 70Yb) with the model of a single cluster of nucleons revolving about a sphere.
Authors: Pauling L
Published: May 2006

Analysis of the first four excited energy levels of the ground-state rotational bands of 82 nuclei (isotopes of the seven even-even lanthanons from 58Ce to 70Yb) has provided values of the radius of revolution of a single cluster about a sphere in each nucleus. The number of nucleons m in each cluster is taken to be 2 for neutron number N = 82 in each case for the first excited state, usually increasing by 2 with change in Z or N by +/-2, but sometimes changing by 0 or by 4 or more. The radius of revolution R decreases to a minimum (about 6.7 fm) at N approximately 78, rises sharply at N approximately 86, reaches a maximum (about 7.8 fm) at N approximately 96, and then decreases.
Active conformation of an insect neuropeptide family.
Authors: Nachman R J , Roberts V A , Dyson H J , Holman G M , Tainer J A
Published: May 2006

To understand the structural and chemical basis for insect neuropeptide activity, we have designed, synthesized, and determined the conformation of a biologically active cyclic analog of the pyrokinins, an insect neuropeptide family that mediates myotropic (visceral muscle contractile) activity. Members of this insect neuropeptide family share the common C-terminal pentapeptide sequence Phe-Xaa-Pro-Arg-Leu-NH2 (Xaa = Ser, Thr, or Val). Circular dichroic, nuclear magnetic resonance, and molecular dynamics analyses of the conformationally restricted cyclic pyrokinin analog cyclo(-Asn-Thr-Ser-Phe-Thr-Pro-Arg-Leu-) indicated the presence of a beta-turn in the active core region encompassing residues Thr-Pro-Arg-Leu. The rigid cyclic analog retains biological activity, suggesting that its C-terminal beta-turn is the active pyrokinin conformation recognized by the myotropic receptor. As individual pyrokinins and pyrokinin-like neuropeptides demonstrate both oviduct-contractile and pheromone-biosynthesis activities in various insects, the biologically active beta-turn structure reported here holds broad significance for many biological processes.