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)

The 5' noncoding region sequence of rabbit beta-globin mRNA has been determined. This region is 53 nucleotides long, not including the A-U-G initiator sequence or m7Gppp "cap" structure. 32P-labeled DNA complementary to the 5' noncoding region was synthesized using reverse transcriptase with the synthetic deoxyoctanucleotide d(G-C-A-C-C-A-T-T) as a "primer". The cDNA produced was then sequenced using a modification of the gel-sequencing technique previously developed for DNA sequencing (G.G. Brownlee and E.M. Cartwright, manuscript in preparation). The sequence obtained was checked by depurination and nearest-neighbor analysis. The known sequence at the 3' end of rabbit 18S ribosomal RNA cannot base-pair extensively with the 5' noncoding region of beta-globin mRNA; however, it does form 6 base pairs around the initiation codon.
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PMID:Complete nucleotide sequence of the 5' noncoding region of rabbit beta-globin mRNA. 6 96

We have determined the terminal heteropolymeric sequences of AMV RNA by the following procedures: first, RNA sequence determination on the 5' terminal and the poly(A)-linked 3' terminal T1 oligonucleotides, and second, analysis by the Maxam and Gilbert (1977) method of AMV strong stop DNA and of DNA complementary to the poly(A)-linked T1 oligonucleotide, synthesized with reverse transcriptase and (pdT)13 as primer. The structure deduced for the 5' terminal region is (5')7mGpppGmCCAUUCUACCUCUCACCACAUUGGUGUGCACCUGGGUUGAUGGCCGGACCGUCGAUUCCCUGACGACUACGAGCACCUGCAUGAAGCAGAAGGCUUCAU... Two distinct 3' terminal sequences were deduced: GCCAUUCUACCUCUCAAA...AOH and GCCAUUCUACCUCUCACCAAA...AOH. The two termini, differing by a C-C-A sequence, may reflect genetic heterogeneity of the AMV stock or, more probably, may be generated at or after RNA transcription. These results demonstrate a terminal redundancy of the hetero polymeric sequence of 16 and 19 nucleotides, respectively. The terminal redundancy allows for mechanisms which involve transfer of the DNA segment synthesized on the 5' terminal redundant sequence to the 3' terminal redundant sequence.
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PMID:Avian myeloblastosis virus RNA is terminally redundant: implications for the mechanism of retrovirus replication. 19 42

We have isolated a cDNA clone for one of the HLA-B locus alloantigens by hybridization with a 30-nucleotide-long DNA probe. The probe was isolated from a reverse transcriptase (RNA-dependent DNA nucleotidyltransferase)-catalyzed cDNA synthesis reaction on poly(A)-mRNA in which an oligonucleotide (5'-32P)dC-T-T-C-T-C-C-A-C-A-TOH served as a primer and in which dideoxynucleoside triphosphates were used to reduce the size and heterogeneity of the cDNA products. The desired cDNA clone was isolated from a library of recombinant cDNA clones in the plasmid pBR322. The partial nucleotide sequence of the cDNA clone corresponds to the amino acid sequence of HLA-B7 antigen. The approach described in this paper is extremely sensitive and may be useful in cloning other genes for which the corresponding mRNA is present at low levels. This cDNA clone is nearly full length and can be used to isolate and to study the genes within the HLA region and to obtain expression of HLA-B peptides in cells.
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PMID:Isolation and partial nucleotide sequence of a cDNA clone for human histocompatibility antigen HLA-B by use of an oligodeoxynucleotide primer. 616 99

Pancreatic poly(A) RNA isolated from the channel catfish (Ictalurus punctatus) was enriched for sequences corresponding to somatostatin mRNA on isokinetic sucrose gradients. Double-stranded cDNA was synthesized and inserted into the Pst I site pBR322 via the poly(dG) . poly(dC) tailing method. Escherichia coli was transformed with this DNA, and colonies containing somatostatin cDNA sequences were identified by hybridization using a primer-extended somatostatin cDNA. The somatostatin cDNA was obtained by extending a 5'-labeled undecanucleotide primer complementary to somatostatin mRNA with reverse transcriptase using catfish poly(A) RNA as a template. The synthetic primer d(T-T-C-C-A-G-A-A-G-A-A) was deduced from the amino acid sequence Phe-Phe-Trp-Lys present in somatostatin-14. Twenty positive colonies were obtained upon screening 2000 transformants. The restriction maps of the plasmid DNA obtained from the positive colonies were examined. Nineteen of these plasmids contained sequences corresponding to somatostatin-14, while one contained a sequence corresponding to somatostatin-22. The nucleotide sequence of pancreatic somatostatin-14 is reported here. The cDNA contains 350 nucleotides in the 3' noncoding region, 345 nucleotides in the coding region, and 104 nucleotides in the 5'-untranslated region. The mRNA codes for a precursor to somatostatin which is 114 amino acids in length. The preprosomatostatin has a sequence of hydrophobic amino acids at the NH2 terminus, followed by a connecting peptide of approximately 75 amino acids. The sequence Arg-Lys precedes somatostatin-14. Analysis of genomic DNA from the channel catfish reveals that somatostatin-14 and somatostatin-22 are present on different restriction fragments.
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PMID:The structure of cloned DNA complementary to catfish pancreatic somatostatin-14 messenger RNA. 617 39

mRNA for bacteriorhodopsin from Halobacterium halobium has been partially purified. By using this mRNA as template in the presence of reverse transcriptase RNA-dependent DNA nucleotidyltransferase and a 5'-[32P] synthetic oligodeoxyribonucleotide corresponding to amino acids 9-12 of bacteriorhodopsin as primer, we have isolated the major 5'-[32P]cDNA product, approximately 80 nucleotides long, and determined its sequence. Based on the cDNA sequence, the 5'-proximal sequence of bacteriorhodopsin mRNA is G-C-A-U-G-U-U-G-G-A-G-U-U-A-U-U-G-C-C-A-A-C-A-G-C-A-G-U-G-G-A-G-G-G-G-G-U-A-U-C -G-C-A-G-G-C-C-C-A-G-A-U-C-A-C-C-G-G-A-C-G-U-C-C-G. This includes the expected sequence for amino acids 1-8 and shows that bacteriorhodopsin is synthesized as a precursor that is at least 13 amino acids longer (Met-Leu-Glu-Leu-Leu-Pro-Thr-Ala-Val-Glu-Gly-Val-Ser) at the NH2 terminus. Agarose/urea gel electrophoresis of the partially purified mRNA showed several bands; of these, a major one hybridized with 5'-[32P]cDNA. These results suggest that the bacteriorhodopsin mRNA in the partially purified preparation is homogeneous in size and that it constitutes a substantial portion of the RNA preparation subjected to electrophoresis.
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PMID:Bacteriorhodopsin: partial sequence of mRNA provides amino acid sequence in the precursor region. 694 48

We have used an oligodeoxynucleotide of defined sequence to detect and quantitate proenkephalin mRNA in the poly(A)-containing fraction of RNA from bovine adrenal medullas. The decahexamer 5'-d(G-G-T-A-G-T-C-C-A-T-C-C-A-C-C-A)-3' was synthesized to be complementary to the codons specifying the amino acid sequence NH2-Trp-Trp-Met-Asp-Tyr-Gln-COOH. This stretch of amino acids occurs in peptide I, one of the intermediates in the biosynthetic pathway of the enkephalins in bovine adrenal medulla. This pathway starts with a precursor (proenkephalin) of about 45 kilodaltons [Stern, A. S., Jones, B. N., Shively, J. E., Stein, S. & Udenfriend, S. (1981) Proc. Natl. Acad. Sci. USA 78, 1962-1966]. The decahexamer hybridized to adrenal poly(A)+RNA and was extended into cDNA with reverse transcriptase (RNA-dependent DNA nucleotidyltransferase). Five main discrete products ranging in size from 115 to 168 nucleotides were observed. The sequences of these extensions were found to be identical over the approximately 70 nucleotides sequenced from their 5' termini and corresponded exactly to the sequence expected from the amino acid sequence of peptide I. These cDNAs and the decahexamer itself hybridized to an adrenal medullary poly(A)+RNA species of about 1500 nucleotides, sufficient in size to code for the proposed proenkephalin. At saturation, approximately 2 fmol of the decahexamer were bound per microgram of mRNA; thus, the proenkephalin mRNA represents about 0.1% of the total poly(A)+RNA population in the tissue.
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PMID:Detection and partial characterization of proenkephalin mRNA. 694 86

The structure of a complementary hybrid duplex of RNA and DNA has been determined by X-ray crystallography. A ten residue DNA oligonucleotide of sequence 5'-G-G-C-G-C-C-C-G-A-A-3' was annealed to complementary RNA (5'-u-u-c-g-g-g-c-g-c-c-3') and crystallized, producing tetragonal crystals that diffract to 2.3 A resolution. The hybrid adopts a geometry that is neither strictly A nor B-form, rather the helix possesses qualities of both, reminiscent of spectroscopic descriptions of a hybrid conformation, or H-form. All of the ribonucleotides maintain the C3'-endo conformation seen in A-form, while both C3'-endo and C2'-endo conformations are found in the deoxyribonucleotides. The minor groove width (8.5 to 10.5 A) is intermediate between standard values for A (11 A) and B-form (7.4 A) DNA. The global parameters rise and base-pairs tilt (or inclination) are like that of A-DNA, however the slide and x displacement (Dx) are more like that of A-RNA, thus giving the hybrid a unique conformation. In addition, the 10-mer crystallizes in a manner that allows the formation of dimers that stack end-to-end, thereby providing a glimpse of how an extended (20 base-pair) helix of RNA-DNA hybrid might appear. This duplex sequence was selected for study because it is specifically recognized by the ribonuclease H function of HIV reverse transcriptase. A structure of a substrate of this enzyme is of potential value in understanding requirements for the selectivity of this important drug target. The minor groove of the hybrid duplex, lined with the 2-OH of the ribose rings, is the single distinguishing characteristic of the RNA/DNA hybrid, undoubtedly an important structural feature conferring selectivity.
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PMID:The structure of an RNA/DNA hybrid: a substrate of the ribonuclease activity of HIV-1 reverse transcriptase. 896 2

Human monocyte chemotactic factor-3 (MCP-3) belongs to the C-C chemokines, which are cytokines involved in cell recruitment in inflammation and cancer. Northern blotting and reverse transcriptase polymerase chain reaction (RT-PCR) analyses showed that the MCP-3 gene is expressed in many human tissues and tumor cell lines and that the expression level is increased by various stimuli. Measles virus and phorbol 12-myristate 13-acetate (PMA) induced MCP-3 mRNA after 6 hr of stimulation. Interferon-beta (IFN-beta) induced MCP-3 mRNA after 16 hr, a time point when the PMA-induced mRNA had the tendency to level off. No significant increase in MCP-3 mRNA levels was observed in MG-63 cells after stimulation with interleukin-1beta (IL-1beta). To elucidate the regulation of MCP-3 gene expression, we determined the sequence of 5 kb of the MCP-3 promoter. This sequence contained a microsatellite that was shown to be polymorphic in various cell lines. Next 5'-deletion mutants of the promoter were generated and transfected into MG-63 cells, demonstrating the presence of several positive and negative transcriptional regulatory elements. One of the positive elements was located at -37, only 21 bp upstream from the TATAA box. This element was similar to an AP-1 element and also to a homeodomain protein Pbx1 binding site. A deletion mutant from -110 to +52 possessed the highest promoter activity, and the longer deletion mutants had relatively low activities. The region between -190 and -172 contained an Ets-like element and inhibited promoter activity. Stimulation with PMA dramatically increased promoter activity through activation of a positive element present between -172 and -100. The same 5'-deletion mutants were transfected into HeLa and Jurkat cells. None of the deletion mutants had any significant activity in Jurkat cells. In HeLa cells, low levels of MCP-3 mRNA were detected by RT-PCR, but the profile of the promoter activities of the deletion mutants was different from that seen in MG-63 cells.
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PMID:Structural and functional analysis of the promoter region of the human MCP-3 gene: transactivation of expression by novel recognition sequences adjacent to the transcription initiation site. 905 38

Infection with human immunodeficiency virus (HIV)-1 most often leads to the development of acquired immune deficiency syndrome, which may manifest with opportunistic infections, many of which occur in the lung. Mononuclear phagocytes infected by HIV-1, being relatively resistant to its cytopathic effects, potentially act as a reservoir for the virus. The alveolar macrophage (AM), a differentiated lung tissue macrophage, is readily infected by HIV-1, after which the virus becomes relatively dormant. C-C chemokines, secreted by CD8 T lymphocytes and other cells, are known to suppress HIV replication in lymphocytes. In view of this observation, and the relative increase in CD8+ T lymphocytes during HIV-1 disease, particularly in the lung, we hypothesized that C-C chemokines might play a key role in suppressing HIV-1 replication in AM. We examined the effect of the C-C chemokines macrophage inflammatory protein (MIP)-1 alpha, MIP-1 beta, and regulated on activation normal T cell expressed and secreted (RANTES) singly and in combination on HIV-1 replication in peripheral blood monocytes (PBM) and AM infected in vitro. Our findings indicate that RANTES suppresses HIV-1 replication, as measured by reverse transcriptase activity, in PBM (41.3 +/- 15.2% of control, n = 3, P < 0.05) and AM (30.3 +/- 7.8% of control, n = 3, P < 0.05) in a dose-dependent manner. The other C-C chemokines had no significant effect singly (MIP-1 alpha PBM: 64.8 +/- 21.9%; AM: 115.0 +/- 2.4% of control; MIP-1 beta PBM: 68 +/- 19.6; AM: 63.3 +/- 26.2% of control) but modestly decreased HIV replication when incubated in addition to RANTES (24.5 +/- 6.5% of control). These observations suggest that RANTES plays a key role in modulating HIV-1 replication in mononuclear phagocytes in the blood and lung, and this may have therapeutic implications for prevention and/or treatment of HIV disease.
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PMID:RANTES inhibits HIV-1 replication in human peripheral blood monocytes and alveolar macrophages. 917 70

The chemokine regulated on activation, normal T cells expressed and secreted (RANTES), is a C-C chemokine and a potent chemoattractant for monocytes, T lymphocytes, basophils, and eosinophils. Its expression by human airway epithelium has been demonstrated both in vitro and in vivo. We investigated whether RANTES is expressed by normal human airway epithelial cells after influenza viral infection and examined its bioactivity. Epithelial cells were obtained from bronchial tissue or nasal polyps of patients who had undergone lobectomy for lung cancer or polypectomy for nasal polyps. These cells were cultured by the outgrowth method. Cultured cells were infected with influenza virus A (subtype H3N2) after which the supernatants and the cells were collected 8 to 72 h after infection. RANTES mRNA (messenger RNA) was analyzed by the reverse transcriptase-polymerase chain reaction and Southern blot analysis of its product. Concentrations of RANTES in the supernatants were analyzed by enzyme-linked immunosorbent assay. RANTES protein and mRNA were not detected in the media of uninfected cells. PCR products for RANTES were clearly detected in nasal and bronchial epithelial cells 24 h after infection. Southern blot analysis confirmed that the PCR products were indeed specific for RANTES mRNA. Twenty-four to 72 h after infection, significant levels of RANTES protein were detected in culture media. We also investigated the chemotactic activity of the supernatant of cultured cells. The supernatant of the cells 48 h after infection had potent chemotactic activity for eosinophils, which was attenuated by the addition of anti-RANTES antibodies. These findings suggest that influenza virus infection may induce expression of bioactive RANTES by normal human bronchial and nasal epithelial cells.
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PMID:Expression of RANTES by normal airway epithelial cells after influenza virus A infection. 947 13


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