Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bovine herpesvirus 1 (BHV-1) establishes a latent infection in sensory ganglionic neurons of infected animals. Expression of latency-related (LR) gene products is controlled by a 980-bp fragment (LR promoter). DNA sequence analysis revealed that two major open reading frames (ORFs) are in the LR gene. Antibodies directed against both ORFs were generated in rabbits by using synthetic peptides. Antibody P2, which is directed to sequences near the amino terminus of ORF 2, recognized a 41-kDa protein in lytically infected cells, suggesting that ORF 2 encodes a protein. When the LR gene was inserted into a mammalian expression vector and subsequently transfected into COS-7 cells, a 41-kDa protein was detected by use of silver-stained sodium dodecyl sulfate-polyacrylamide gels and by the P2 antibody. In contrast, this protein was not detected in mock-transfected cells. Deletion of DNA sequences containing ORF 2 blocked synthesis of the 41-kDa protein in COS-7 cells. Reverse transcriptase-mediated PCRs indicated that splicing occurs near the C terminus of ORF 2. Further studies indicated that LR RNA was alternatively spliced in latently infected cattle and that a fraction of LR RNA was poly(A)+. Taken together, these studies suggested that a spliced LR transcript has the potential to encode a 41-kDa protein.
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PMID:Identification of gene products encoded by the latency-related gene of bovine herpesvirus 1. 763 78

The L-type voltage-dependent calcium channel (L-VDCC) is assumed to be a critical component of excitation-contraction coupling in smooth muscle. Using pregnant rat myometrium, we examined the hypothesis that parturition is associated with significant changes in the expression of the alpha 1 subunit of the L-VDCC at the mRNA or protein level. The binding of radiolabeled dihydropyridine, which correlates with the total number of calcium channels in the membrane, was increased by 14 days' gestation, in comparison to that in nonpregnant controls. The elevation in binding capacity persisted through labor and fell postpartum. Northern and RNA dot-blot analysis demonstrated the highest level of expression on Days 20 and 21, with a 3- to 10-fold decrease during parturition. We believe these studies are most consistent with a one-day lag time between mRNA and protein expression, and generally support a modest increase in L-VDCC expression in pregnancy and labor. Reverse transcriptase polymerase chain reaction was used to examine changes in isoform expression in Motif IV, a region of the alpha 1 subunit known to be alternatively spliced. These studies revealed the presence of multiple isoforms in rat myometrium, with a predominance of IVS3B. Interestingly, a marked increase in the ratio of S3B:S3A was noted at parturition. In summary, these data demonstrate that the number of L-type calcium channels, although increased in pregnancy, do not change prior to, or with the onset of, myometrial contraction. Intriguingly, mRNA expression was markedly decreased at parturition. The change in isoform expression during labor is of unknown, but potential, physiologic significance.
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PMID:Changes in the expression of the L-type voltage-dependent calcium channel during pregnancy and parturition in the rat. 784

Examination of cDNAs for the laminin-binding alpha 7 integrin subunit identified two different sequences (designated X1 and X2) coding for the variable region between the III and IV homology repeat domains near the putative ligand-binding site. Sequencing of a mouse alpha 7 genomic clone established that the X1 and X2 regions are derived by mutually exclusive alternative mRNA splicing. Reverse transcriptase-polymerase chain reaction analysis of alpha 7 mRNA indicated that the X1 and X2 isoforms were present in equal amounts in mouse skeletal myoblasts and adult heart. However, in adult skeletal muscle, the X2 variant was exclusively expressed. Amino acid sequence homologies in the III/IV segment suggest that alpha 3 and alpha 6 are also alternatively spliced at this site. We identified alternatively spliced exons in a human alpha 6 genomic clone that encode X1- and X2-like segments. Analysis of the alpha 7 cytoplasmic domain indicated that this region was also alternatively spliced and like alpha 3 and alpha 6 could exist as the A or B form. In mouse skeletal and cardiac muscle the B form of alpha 7 was strongly expressed. However, we identified alpha 7A in neonate and adult skeletal muscle but not in cardiac tissue. High levels of alpha 7A were detected in differentiating myotubes, but in proliferating myoblasts only the alpha 7B isoform was present. These results indicate that alternative splicing of alpha 7 mRNA is differentially regulated during development and generates variant integrin chains with structurally and presumably functionally unique ligand-binding and cytoplasmic domains.
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PMID:Alternative extracellular and cytoplasmic domains of the integrin alpha 7 subunit are differentially expressed during development. 825 14

In the rabbit heart, multiple isoforms of cardiac troponin T (cTnT1 through cTnT5, from largest in size to smallest), a protein essential for calcium-regulated myofibrillar ATPase activity, have been identified, and a correlation has been found between these isoforms and myofilament sensitivity to calcium. We have sought to establish the molecular basis of this diversity. Restriction-digest analysis of genomic DNA has indicated that the rabbit cTnT gene is a single-copy gene. cTnT cDNA clones were isolated from cDNA libraries, yielding a consensus sequence for the protein. Newborn rabbit heart cDNAs, obtained using the reverse-transcriptase polymerase chain reaction (RT-PCR), were amplified using primers derived from this cDNA. Three full-length cDNAs that differed by the inclusion or exclusion of three short nucleotide sequences within the cDNAs were obtained. Amplification in the 5' half of the cDNAs confirmed that multiple cTnT products arose because of the variable inclusion of an 18- and a 30-nt sequence. The 30-nt sequence has homology with previously described alternatively spliced exons in rat and chicken cTnT, whereas the 18-nt sequence has not been described previously. RT-PCR in the 3' half of the cDNAs confirmed an additional region of heterogeneity: the presence, in part or in full, or absence of a 9-nt region, which matches the alternatively spliced exon 12 described for rat cTnT. In vitro transcription and translation of four cDNA clones containing both the 18- and 30-nt sequences, the 30-nt sequence, the 18-nt sequence, or neither generated protein isoforms that comigrated with cTnT1, cTnT2, cTnT3, and cTnT4, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Molecular basis of cardiac troponin T isoform heterogeneity in rabbit heart. 826 93

Sequence analysis has shown significant homology between the catalytic regions of the mammalian ras GTPase-activating protein (GAP), yeast Ira1p and Ira2p (inhibitory regulators of the RAS-cyclic AMP pathway), and neurofibromin, the protein encoded by the NF1 gene. Yeast expression experiments have confirmed that a 381-amino-acid segment of neurofibromin, dubbed the GAP-related domain (GRD), can function as a GAP. Using the RNA polymerase chain reaction with primers flanking the NF1-GRD, we have identified evidence for alternative splicing in this region of the NF1 gene. In addition to the already published sequence (type I), an alternative RNA carrying a 63-nucleotide insertion (type II) is present in all tissues examined, although the relative amounts of types I and II vary. The insertion is conserved across species but is not present in GAP, IRA1, or IRA2. GenBank searches have failed to identify significant similarity between the inserted sequence and known DNA or protein sequences, although the basic amino acid composition of the insertion shares features with nuclear targeting sequences. Expression studies in yeasts show that despite the partial disruption of the neurofibromin-IRA-GAP homology by this insertion, both forms of the NF1-GRD can complement loss of IRA function. In vivo assays designed to compare the GAP activity of the two alternatively spliced forms of the NF1-GRD show that both can increase the conversion of GTP-bound ras to its GDP-bound form, although the insertion of the 21 amino acids weakens this effect. The strong conservation of this alternative splicing suggests that both type I and II isoforms mediate important biological functions of neurofibromin.
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PMID:A conserved alternative splice in the von Recklinghausen neurofibromatosis (NF1) gene produces two neurofibromin isoforms, both of which have GTPase-activating protein activity. 841 46

The C57BL/6 mouse differs from the BALB/c mouse in that its ear and skin mast cells and its progenitor bone marrow-derived mast cells (mBMMCs) do not express mouse mast cell protease (mMCP) 7. We now report that, as detected by nuclear run-on analysis, the mMCP-7 gene is transcribed in C57BL/6 mBMMCs at a rate comparable to that in BALB/c mBMMCs. Reverse transcriptase-polymerase chain reaction analysis and sequencing of the product revealed that the ears of C57BL/6 mice contain small amounts of a mMCP-7 transcript that possesses a 98-base pair deletion. The deletion begins at a normally quiescent cryptic splice site (G416TGAG), 98 base pairs upstream of the normal exon 2/intron 2 splice site (G514TGAG), and introduces a premature stop codon in the alternatively spliced transcript. Thus, even if translated, the mature protein would consist of only 18 amino acids as compared to 245 amino acids in normal mMCP-7. Sequence analysis of the mMCP-7 gene in the C57BL/6 mouse revealed that the cryptic splice site is activated due to a G514-->A point mutation at the first nucleotide of the normal exon 2/intron 2 splice site. This is the first report of a mutation of a gene that encodes a mast cell secretory granule constituent that leads to its loss of expression. Moreover, the mMCP-7 gene is the first found in any species that sequentially has undergone a splice site mutation to cause retention of an intron and then a second splice site mutation to cause activation of a cryptic splice site.
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PMID:Natural disruption of the mouse mast cell protease 7 gene in the C57BL/6 mouse. 857 65

The adhesion molecule intercellular adhesion molecule-1 (ICAM-1), in addition to its membrane-associated form (mICAM-1), also exists as a soluble form (sICAM-1). sICAM-1 is capable of binding to lymphocyte function associated antigen-1 (LFA-1) molecules, and production of sICAM-1 is therefore thought to have immunomodulatory consequences. The present study, which employed normal human keratinocytes as a model for sICAM-1-producing cells, was conducted to determine the mechanism responsible for the production of sICAM-1 and to develop a strategy for specific inhibition of sICAM-1 production. Stimulation of keratinocytes with recombinant human gamma-interferon (rhIFN-gamma) induced both expression of mICAM-1 and production of sICAM-1. Western blot analysis revealed that keratinocyte-derived sICAM-1, compared to mICAM-1, had a smaller molecular size, approximately a 7-kD difference. Neither by Northern blot analysis nor by reverse-transcriptase polymerase chain reaction (RT-PCR) was any evidence for alternatively spliced ICAM-1 mRNA obtained. Addition of the protease inhibitors iodoacetamide and E-64, however, inhibited the production of sICAM-1 in a dose-dependent manner. The involvement of proteolytic cleavage in the production of sICAM-1 was corroborated in minimal peptide protection assays, in which minimal peptides covering the potential cleavage site of ICAM-1 were added to sICAM-1-producing keratinocytes. One of these peptides, ICAM cleavage inhibitory peptide (ICAM-CIP), inhibited the production of sICAM-1 without affecting mICAM-1 expression. These studies demonstrate that sICAM-1 production in human keratinocytes is due to proteolytic cleavage, and that the oligopeptide ICAM-CIP may specifically inhibit this mechanism. The capacity of ICAM-CIP to selectively prevent production of sICAM-1 may be useful for the development of novel therapeutic approaches relevant for the management of inflammation and cancer.
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PMID:Analysis of the production of soluble ICAM-1 molecules by human cells. 864 65

Four different plasma membrane Ca(2+)-ATPase (PMCA) genes and three sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) genes have been previously cloned and characterized. In this study we have investigated the expression of the mRNA encoding the various PMCA and SERCA proteins in fetal and adult human heart and placenta by the reverse-transcriptase-polymerase-chain-reaction (RT-PCR) and cDNA cloning. We have found that PMCA1 and PMCA4 genes were expressed in 8-, 12- and 20-week fetal heart and in adult heart. PMCA2 gene was expressed at low levels in adult heart but was not detected in fetal heart. PMCA3 mRNA was not detected in the heart nor placenta. In contrast, the mRNA encoding SERCA2a, SERCA2b and SERCA3 were expressed in all cardiac developmental stages. Multiple alternatively spliced mRNA transcripts which differ at splice site A and B/C of the PMCA1, PMCA2 and PMCA4 genes were detected in the human heart. Interestingly, a novel tissue specific variant of the PMCA4 gene was detected in both fetal and adult human heart but not in placenta that accounts for about 30% of the total PMCA4 mRNA variant expression. DNA sequence analysis of this novel variant revealed that it corresponds to the equivalent of the PMCA1d variant and accordingly we have named it PMCA4d. We cloned and sequenced eight cDNA inserts encoding for the PMCA1 and PMCA4 variants from a fetal human heart cDNA library confirming that these are the two main PMCA genes expressed in cardiac muscle.
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PMID:Analysis of mRNA expression and cloning of a novel plasma membrane Ca(2+)-ATPase splice variant in human heart. 870 Jan 62

The prostate gland contains neuroendocrine cells and amidated neuroendocrine peptides whose presence has been related to aggressive forms of prostate cancer. The enzyme peptidylglycine alpha-amidating monooxygenase (PAM) is critical to the bio-synthesis of amidated peptides and is commonly present in neuroendocrine cells. By northern blot hybridization analysis, PAM mRNA was detected in similar quantities in dorsolateral and ventral prostates of 3-month-old and 13-month-old rats. Multiple forms of PAM mRNA were present whose size distribution was more similar to PAM mRNAs found in pituitary than atrium. Alternative splice sites in PAM mRNA were investigated by reverse-transcriptase polymerase chain reaction. Similar alternatively spliced forms of PAM mRNA were found in both prostate lobes, pituitary, and atrium. However, the distribution of forms in the prostate most resembled that of pituitary. Multiple forms of PAM mRNA are present in prostate and may serve as markers of neuroendocrine differentiation.
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PMID:Expression and processing of peptidylglycine alpha-amidating monooxygenase messenger RNA in rat prostate. 870 Jul 99

Myelin/oligodendrocyte glycoprotein (MOG) is an integral membrane protein expressed on the oligodendrocyte cell surface and the outermost surface of myelin sheaths. Due to this localization, MOG is a primary target antigen involved in immune-mediated demyelination. We previously reported that MOG is a unique member of the immunoglobulin (Ig) superfamily in that it possesses two large hydrophobic domains. MOG is highly conserved between deduced peptide sequences of rodent and human MOG (approximately 89% identity). We have completed an investigation of alternative splicing within the human and mouse MOG genes. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of total cellular RNA isolated from both fetal and adult human central nervous system (CNS) tissues reveals a complex array of alternatively spliced MOG-specific variants and the presence of two novel exons. Exon 3 encodes a short hydrophilic domain containing multiple in-frame termination codons that would result in truncation of MOG prior to translation of its transmembrane domain. Exon 7 encodes an additional hydrophilic domain that replaces MOG's second hydrophobic domain in one splice variant. We also observed that five of our eight MOG variants exhibited an alternative internal 3' splice acceptor within MOG's terminal exon. Surprisingly, no splicing was observed in a developmental study using mouse brainstem RNA.
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PMID:Myelin/oligodendrocyte glycoprotein is alternatively spliced in humans but not mice. 891 5


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