Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.49 (reverse transcriptase)
 31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A cell line, TBSV7, that produces noninfectious murine sarcoma virus (MuSV) in the absence of helper MuLV was isolated from TB cells infected with the supernatant of MuSV349 cells. These noninfectious MuSV particles with "immature" C-type virus morphology contain a 2.2 X 10(6)-Da genomic RNA and an uncleaved 62,000-Da gag precursor protein (Pr62). Neither viral envelope proteins (gp70, p15E, p12E) nor reverse transcriptase were detected in these virus particles. Pr62 was found to be phosphorylated in vivo and it could be phosphorylated in vitro with [gamma-32P]ATP, indicating that protein kinase was packaged in these noninfectious virions. In vitro processing of Pr62 to smaller molecular weight proteins could be achieved by the addition of Mo-MuLV and Nonidet P-40. The initial cleavage products were proteins with molecular weights of 38K (Pr38) and 27K (Pr27). Under optimum conditions Pr38 was cleaved to p30 and a protein band migrating with MuLV-p10, while Pr27 was cleaved to a 17,000-Da protein that migrated slower than MuLV-p15 and a protein band migrating with MuLV-p12. Pulse-chase experiments performed on TBSV7 cells superinfected with Mo-MuLV indicated that intracellular processing of Pr62 was much slower than that of Pr65. Cleavage protein products of Pr62 similar in size to the in vitro protein products were also detected in TBSV7 cells superinfected with MuLV.
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PMID:Isolation and characterization of a Mo-MuSV-transformed TB cell line that produces noninfectious MuSV particles with uncleaved gag protein which is processed in the presence of Mo-MuLV. 632 21

A new cell line designated SQ-A was established from the spleen of a leukemic DBA/2J mouse inoculated with the anemic strain of Friend erythroleukemia virus (FLV-A). The cells are similar in morphology, growth pattern, and tumorigenicity to our prototype erythroleukemia line 5-86 but are more sensitive to the cytotoxic effects of inducers of differentiation. The virus produced by SQ-A cells induces erythroleukemia associated with anemia in adult mice but has little activity when assayed on XC cells. It was characterized to determine what factors influence its leukemogenic potential. As compared to the attenuated virus from cultures of 5-86 and G-2 cells, the subunits of the RNA from the virions of SQ-A cells are the same size, and the amount of reverse transcriptase activity and RNase H present in the purified virions of the three lines are similar. However, differences are observed in levels of endonuclease and protein kinase. Both enzymes are increased in SQ-A virions. The activity of protein kinase in SQ-A virions is about 5 times higher than that in the attenuated virions. The number of polypeptides and their phosphorylation patterns also distinguish the virions of SQ-A. Whereas 5-86 virions contain seven proteins, three of which are phosphorylated in vitro, SQ-A virions contain eight proteins, all of which are phosphorylated. The extra protein in SQ-A virions has a molecular weight of 25,000 and is not glycosylated.
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PMID:Characterization of leukemogenic virus produced by a new line of Friend erythroleukemia virus-transformed cells. 632 17

Retroviruses are RNA-containing viruses using reverse transcriptase to produce DNA copies capable of insertion into host chromosomes. Appropriate genes are required to confer transforming ability to retroviruses. The src gene, a 60,000-dalton protein with protein kinase activity, is required by avian viruses to induce sarcomas. Normal cells have a gene (sarc) similar to the src gene. Retroviruses with oncogenic properties can arise by recombining with genes on the host chromosome. Herpesviruses, adenoviruses, and papovaviruses have transforming properties residing in only a portion of the genome. Probably, only one to two genes are required for transformation, regardless of the complexity of the virus.
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PMID:Current knowledge of mechanisms of viral carcinogenesis. 638 Sep 62

We describe a human (h) PRL-producing cell line, SKUT-1B-20, which we isolated as a subclone of a uterine sarcoma cell line. Although this cell line is of uterine origin, it does not use the decidual-specific upstream promoter of the hPRL gene, but transcribes the hPRL gene from the downstream pituitary-type transcription start site, as determined by Northern blot, reverse transcriptase-polymerase chain reaction and primer extension analyses. This is particularly intriguing because SKUT-1B-20 cells lack the transcription factor Pit-1. No Pit-1 messenger RNA was detectable by reverse transcriptase-polymerase chain reaction, and endogenous Pit-1 target genes (GH, PRL, and Pit-1) were refractory to transfected Pit-1 expression vector, whereas in cotransfection experiments, Pit-1 efficiently activated reporter gene fusion constructs carrying 5'-flanking sequences of the human and rat PRL or the mouse Pit-1 genes. By transfecting reporter genes containing 8.7 kilobases of DNA flanking the hPRL pituitary-specific start site (hPRL-8700/Luc) and deletions thereof, we located a Pit-1-independent cis-active region more than 7 kilobases upstream of the start site. The most distal 1650 or 880 base pairs of the hPRL genomic fragment (which extends to -8784 base pairs), when placed directly upstream of the homologous hPRL or the heterologous thymidine kinase promoters, conferred transcriptional activation to those promoters. SKUT-1B-20 cell-specific activation of hPRL-8700/Luc could not be suppressed by the introduction of an inhibitor of protein kinase A (PKA), PKI. This is the first demonstration of pituitary-type PRL gene transcription independent of Pit-1 and activation of the PKA pathway. The SKUT-1B-20 cell line was then used in reconstitution experiments to delineate the role of Pit-1 in modulating the transcriptional effects of phorbol ester, PKA, and estrogen receptor (ER) on the hPRL gene. The low response of hPRL/luciferase fusion genes to phorbol ester was greatly enhanced by cotransfected Pit-1 and was mediated by the proximal region between -250 and -38. The catalytic subunit of PKA, C beta, was able to elicit a moderate induction of hPRL-8700/Luc even in the absence of Pit-1. A potential estrogen response element has been located in the hPRL gene sequence at a position similar to that of the estrogen response element of the rat PRL gene immediately adjacent to the distal enhancer.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Pituitary-type transcription of the human prolactin gene in the absence of Pit-1. 747 71

Currently available evidence supports the hypothesis that insulin-like growth factor-I (IGF-I) secreted by small preantral follicles may be involved in stimulating the initial differentiation of the theca interna and, in particular, expression of the LH receptor in pre-theca cells. To test this hypothesis, we examined the effects of IGF-I on LH receptor mRNA expression in theca-interstitial cells (TIC) isolated from the ovaries of hypophysectomized immature rats by percoll gradient centrifugation. TIC (3.5 x 10(4) viable cells/well) were cultured up to 6 days with and without LH (0-10 ng/ml) and IGF-I (0-100 ng/ml). Androsterone in the medium was measured by RIA, and LH receptor mRNA was measured by specific reverse transcriptase-polymerase chain reaction assay. LH receptor mRNA was low in control (untreated) TIC. IGF-I stimulated a dose-related increase (2-fold) in LH receptor mRNA at 2 days (ED50 = 9.0 +/- 1.9 ng/ml) that remained constant at 4 days and then declined to basal levels at 6 days. LH stimulated a dose-related (ED50 = 17.6 +/- 1.0 pg/ml) increase in LH receptor mRNA that reached a maximum of 4-fold at 2 days. At 4 days, LH down-regulated LH receptor mRNA below basal levels, and it had no effect at 6 days. Addition of IGF-I (30 ng/ml) to LH-treated TIC abolished the stimulatory effect of LH throughout the culture period. LH receptor mRNA was highly sensitive to LH since the ED50 was approximately 2.5-fold lower than for stimulation of androsterone production (39.8 +/- 3.8 pg/ml). To understand the molecular mechanism of the synergistic stimulation of androgen production by IGF-I and LH, the effects of IGF-I on the cAMP/protein kinase A (PKA) signaling pathway were examined. When freshly isolated TIC were challenged with IGF-I alone (30 ng/ml), there was no effect on cAMP production or PKA activity, but IGF-I augmented LH stimulation of cAMP production slightly at high concentrations of LH and blocked stimulation of PKA activity by a saturating concentration of LH (3 ng/ml), suggesting that IGF-I increased LH down-regulation of PKA. We next examined the effects of IGF-I on LH receptor number. When TIC were placed into culture, LH/hCG binding sites decreased to approximately 35% of the initial number at 24 h and 25% at 2 days. This decrease was accompanied by a similar loss of cholera toxin- and hCG-stimulated cAMP production.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Insulin-like growth factor-I regulation of luteinizing hormone (LH) receptor messenger ribonucleic acid expression and LH-stimulated signal transduction in rat ovarian theca-interstitial cells. 752 76

The present study reports changes in saliva composition from the rat parotid gland in response to single and repeated administration of epidermal growth factor (EGF). Treatment of rats with EGF (10 micrograms/kg, i.p., twice daily for 3 days) caused an increase in amylase activity in saliva collected from cannulated parotid duct, following stimulation of secretion with pilocarpine, with a corresponding decrease in enzyme activity in the gland. Analysis of parotid gland RNA by reverse transcriptase-PCR generated a single predicted amylase-derived cDNA product of 576 bp. The steady-state levels of mRNA for amylase from EGF-treated parotid total RNA showed a 1.8-fold increase compared to untreated controls. A single dose of EGF (15 min following i.p. injection) elicited an activation of both protein kinase A and protein kinase C activities. While the activation of protein kinase A was still maintained under the chronic EGF regimen, the activity levels of protein kinase C showed down-regulation to untreated control values.
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PMID:Effect of EGF on rat parotid gland secretory function. 753 11

Although the biochemical properties of soluble guanylate cyclase (sGC) have been extensively studied, little is known about the regulation of gene expression of sGC subunits by second messengers. cAMP analogues and elevating agents have been previously shown to alter gene expression in vascular cells. The aim of the present study was to investigate the effects of cAMP-elevating agents on sodium nitroprusside-stimulated sGC activity and to correlate activity changes with mRNA and protein levels in cultured rat aortic smooth muscle cells. Pretreatment of cells with 50 to 1000 mumol/L isobutylmethyl-xanthine or 0.01 to 10 mumol/L forskolin led to a time- and concentration-dependent decrease in sodium nitroprusside-induced cGMP accumulation, first evident after 3 hours of pretreatment with forskolin and 6 hours of pretreatment with isobutylmethylxanthine. Incubation of cells with a protein kinase A-selective inhibitor (H89 or KT 5720) partially or fully prevented the downregulation in sodium nitroprusside-induced cGMP accumulation caused by cAMP-elevating agents. Quantification of reverse transcriptase-polymerase chain reaction products by high-performance liquid chromatography revealed that mRNA for both alpha1- and beta1-subunits of sGC were decreased in cells pretreated with isobutylmethylxanthine and forskolin but not with dideoxyforskolin (inactive analogue). Moreover, protein levels for the sGC alpha1 subunit of cells pretreated with isobutylmethylxanthine and forskolin but not with dideoxyforskolin were decreased as indicated by Western blot analysis. These data indicate that cAMP-elevating agents decrease sGC activity, possibly by decreasing mRNA or protein levels or both.
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PMID:Regulation of vascular smooth muscle soluble guanylate cyclase activity, mRNA, and protein levels by cAMP-elevating agents. 755 33

Human CD6 is a monomeric 105/130-kDa T cell surface glycoprotein that is involved in T cell activation. The apparent discrepancy between the size of the cytoplasmic domain in human (44 amino acids) and mouse (243 amino acids) CD6, led us to use reverse transcriptase-polymerase chain reaction of human peripheral blood lymphocyte mRNA to isolate cDNA clones that include the carboxyl-terminal coding region of human CD6. The nucleotide sequence of the longest human cDNA clone, CD6-PB1, predicts a protein of 668 amino acids with a 244-amino acid cytoplasmic domain similar in size to and possessing 71.5% amino acid sequence identity with the cytoplasmic domain of mouse CD6. This previously unrecognized 244-amino acid cytoplasmic domain does not have significant homology to any other known protein (except mouse CD6), but does possess two proline-rich motifs containing the SH3 domain-binding consensus sequence, a serine-threonine-rich motif repeated three times, three protein kinase C phosphorylation-site motifs, and 10 casein kinase-2 phosphorylation-site motifs. These sequences are likely to play a role in the ability of CD6-specific monoclonal antibodies to stimulate T cell proliferation. Full-length CD6 cDNA containing this cytoplasmic domain sequence encodes a monomeric 105/130-kDa protein that can be immunoprecipitated from the surface of transfected cells and comigrates upon SDS-PAGE with wild-type CD6 immunoprecipitated from PBL. We also isolated two alternatively spliced forms of human CD6 cDNA lacking sequences encoding membrane-proximal regions of the cytoplasmic domain which maintain the same reading frame as CD6-PB1. The short cytoplasmic domain of the previously reported human CD6-15 cDNA clone results from a deletion of a 20-bp segment through use of an alternative 3' splice site, resulting in a frame shift and premature termination of translation relative to the clones we have isolated. These data demonstrate that human CD6 possesses a large cytoplasmic domain containing sequence motifs that are likely to be involved in signal transduction upon stimulation of T cells through CD6 ligation.
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PMID:Human CD6 possesses a large, alternatively spliced cytoplasmic domain. 758 69

Upon withdrawal of interleukin-3 (IL-3) from human factor-dependent erythroleukemic cell line TF-1, bcl-2 mRNA and protein levels decrease within 8 to 24 hours. Accompanying this decrease is the onset of apoptosis as determined by flow cytometric analysis of DNA degradation. By 8 to 18 hours of deprivation approximately 70% to 80% of the cells have entered apoptosis. Downregulation of protein kinase (PK) by a 24-hour incubation in 100 nmol/L 12-O-tetradecanoyl-phorbol-13-acetate (TPA) in the presence of IL-3 dramatically reduced bcl-2 mRNA levels, and induced apoptosis in the presence of IL-3. We have also found that even in the presence of IL-3, two inhibitors of PKC, light-activated calphostin and H-7, substantially reduced the levels of bcl-2 mRNA between 8 and 24 hours as measured by a semi-quantitative reverse transcriptase/polymerase chain reaction assay method; however, the cyclic nucleotide-dependent PK inhibitor HA 1004, that is a structural analog of H-7 but a poor inhibitor of PKC, did not reduce bcl-2 levels in the presence of IL-3. This decrease in bcl-2 mRNA was accompanied by a decline in bcl-2 protein levels by 8 to 24 hours after addition of light-activated calphostin. In addition to interfering with the maintenance of bcl-2 mRNA levels, inhibition of PKC with H-7 inhibited the induction of bcl-2 mRNA in factor-deprived TF-1 cells restimulated with IL-3. The cyclic nucleotide-dependent PK inhibitor HA 1004 did not inhibit IL-3-induced bcl-2 mRNA. Studies with actinomycin D showed that transcription plays a major role in maintaining bcl-2 levels in TF-1 cells, and it is therefore likely that IL-3 plays a role in maintaining bcl-2 transcription through activation of PKC in these cells.
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PMID:Human interleukin-3 receptor modulates bcl-2 mRNA and protein levels through protein kinase C in TF-1 cells. 779 58

In Drosophila and Caenorhabditis, signal transduction pathways initiated by the activation of receptor-protein tyrosine kinases can mediate developmental fate decisions. In order to examine whether similar mechanisms are employed during mammalian embryogenesis, we undertook a search for novel protein kinases expressed during heart development in the mouse. The primitive mouse heart is formed between 7.75 and 8.5 days post coitum (dpc) and consists of myocardial and endocardial cells. A reverse transcriptase polymerase chain reaction-based approach was used to amplify protein kinase specific products from cDNAs obtained from 8.5 dpc heart tissue. Twenty independent PCR products corresponding to either protein serine/threonine or tyrosine kinases were identified. In this report, we describe the characterization of two of the genes corresponding to the novel PCR products (designated Hek2 and msk). Hek2 encodes the mouse ortholog of human HEK2, a recently identified member of the eph receptor-protein tyrosine kinase gene family. Prior to and at the time of heart formation (7.5-8.0 dpc), Hek2 is expressed in the cranial (rostral) region of the embryo from which a subpopulation of cells will give rise to the rudimentary heart. Between 8.0 and 9.5 dpc, Hek2 mRNA expression is observed in myocardial cells, head mesenchyme and paraxial mesoderm. Hek2 transcripts are not detected in endocardial cells. After 9.5 dpc, Hek2 expression is downregulated. msk (for myocardial SNF1-like kinase) encodes a putative protein serine/threonine kinase most similar to the yeast gene SNF1. msk mRNA expression is restricted to myocardial cells and their progenitors in the 7.75-8.5 dpc developing heart. Subsequently, msk mRNA expression is rapidly downregulated. The patterns of Hek2 and msk expression suggest that these protein kinases may function during development of the primitive heart.
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PMID:Identification of novel protein kinases expressed in the myocardium of the developing mouse heart. 789 99


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