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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In addition to its inhibitory activity against viral DNA polymerases and reverse transcriptase, the acyclic nucleoside phosphonate 9-(2-phosphonylmethoxyethyl)adenine (PMEA) also markedly inhibits the replicative cellular DNA polymerases alpha, delta, and epsilon. We have previously shown that PMEA is a strong inducer of differentiation in several in vitro tumor cell models and has marked antitumor potential in vivo. To elucidate the molecular mechanism of the differentiation-inducing activity of PMEA, we have now investigated the effects of the drug on cell proliferation and differentiation, cell cycle regulation, and oncogene expression in the human erythroleukemia K562 cell line. Terminal, irreversible erythroid differentiation of PMEA-treated K562 cells was evidenced by hemoglobin production, increased expression of glycophorin A on the K562 cell membrane, and induction of acetylcholinesterase activity. After exposure to PMEA, K562 cell cultures displayed a marked retardation of S-phase progression, leading to a severe perturbation of the normal cell cycle distribution pattern. Whereas no substantial changes in c-myc mRNA levels and p21, PCNA, cdc2, and CDK2 protein levels were noted in PMEA-treated K562 cells, there was a marked accumulation of cyclin A and, most strikingly, cyclins E and B1. A similar picture of cell cycle deregulation was also observed in PMEA-exposed human myeloid THP-1 cells. However, in contrast to the strong differentiation-inducing activity of PMEA in K562 cells, the drug completely failed to induce monocytic maturation of human myeloid THP-1 cells. On the contrary, THP-1 cells underwent apoptotic cell death in the presence of PMEA, as demonstrated by prelytic, intracellular DNA fragmentation and the binding of annexin V to the cell surface. We hypothesize that, depending on the nature of the tumor cell line, PMEA triggers a process of either differentiation or apoptosis by the uncoupling of normally integrated cell cycle processes through inhibition of DNA replication during the S phase.
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PMID:9-(2-Phosphonylmethoxyethyl)adenine induces tumor cell differentiation or cell death by blocking cell cycle progression through the S phase. 1039 5

The TCR structure of T cell hybridomas recognizing a tumor glycan-defined epitope has been studied using reverse transcriptase-PCR and gene sequencing. The hybridomas had been raised against a glycopeptide, T72(Tn), consisting of the mouse hemoglobin-derived decapeptide Hb(67 - 76), O-glycoslated in position 72 with alpha-D-GalNAc. The glycan-specific hybridomas varied widely in their use of Valpha genes although Valpha4 was predominant, being present in one third of them. The Vbeta gene usage was more restricted and dominated by Vbeta1 and Vbeta15. There was no correlation between Valpha and Vbeta usage and antigen fine specificity of the hybridomas. The overall amino acid composition of the complementarity-determining region (CDR) 3 of the hybridomas was dominated by small polar residues such as Gly, Asn, Ser, Glu and Ala, amino acids reported in the literature to be frequent in glycan-recognizing proteins. Furthermore, the CDR3 of most hybridomas also contained an aromatic residue with preference for Tyr. A few of the hybridomas raised against the T72(Tn) glycopeptide were peptide specific, i. e. they responded to the unglycosylated peptide only. The amino acid usage of their CDR3 regions was not radically different from that of the glycopeptide specific hybridomas. They also preferentially used Valpha4. However, Vbeta4 and Vbeta8 were the dominating beta chains.
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PMID:Shared structural motifs in TCR of glycopeptide-recognizing T cell hybridomas. 1050 50

The tropical clam Lucina pectinata contains a unique hemoglobin (HbI) which serves to transport H2S to autotrophic bacteria. The cDNA-derived amino acid sequence was obtained from overlapping clones containing the cDNA that codes for HbI. The reverse transcriptase-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) methods were employed to synthesize the cDNA fragments. An initial 354-bp cDNA clone encoding 118 amino acid residues of HbI was amplified from total RNA by RT-PCR using degenerate oligonucleotides. Gene-specific primers derived from the HbI-partial cDNA sequence were used for obtaining the 5' and 3' ends of the cDNA by RACE. The length of the HbI cDNA, estimated from sequence analysis of overlapping clones, was 1322 bp for the full-length cDNA. The coding region of the full-length cDNA codes for 143 amino acid residues. The most conserved amino acid residues in HbI from Lucina pectinata were identified by a multiple alignment with nonvertebrate globin sequences.
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PMID:The cDNA-derived amino acid sequence of hemoglobin I from Lucina pectinata. 1083 19

Globin genes of the bivalve mollusk Scapharca inaequivalvis have the two intron/three exon organization typical of vertebrate and many invertebrate globins, with introns in highly conserved positions. Sequence studies on the A and B globin genes of the mollusk tetrameric hemoglobin gave evidence for the existence of 'minigenes' spanning part of the first and second intron, in-frame with the heme binding domain encoded by the central exon. Putative promoter and regulatory sequences flanking these minigenes were identified in the 3' regions of intron I. Here we report cloning and functional analysis of these regions ( approximately 400bp) and their promoter activity, which was assessed in K562 cells by transient transfection, was established. Moreover, in vitro reverse transcriptase-polymerase chain reaction (RT-PCR) on total cytoplasmatic RNA demonstrated that the A and B minigenes are transcriptionally active in the erythrocytes of S. inaequivalvis. Thus, the present results lead to the conclusion that the present-day organization of the globin genes of S. inaequivalvis tetrameric hemoglobin is still reminiscent of an ancestral globin gene before exon shuffling.
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PMID:Scapharca inaequivalvis A and B miniglobin genes: promoter activity of the 5' flanking regions and in vivo transcription. 1097 67

Suppression subtractive hybridization analysis in our laboratory recently revealed that transferrin mRNA may be elevated in Sedeficient rat liver. In this work, we compared expression in rat liver of genes for transferrin, transferrin receptor, ferritin light and heavy chains, and iron-regulatory proteins 1 and 2 in Se adequacy and deficiency. Weanling male Sprague-Dawley rats were fed Torula yeast diets supplemented with 0 or 0.15 microg Se/kg diet as sodium selenite for 15 wk. Activity of cellular glutathione peroxidase was virtually abolished in Se-deficient rat liver, whereas activity of glutathione S-transferase was 43% higher than in Se-adequate liver. There were no differences in hematocrit, hemoglobin, or liver iron content. To examine differential gene expression, we used a multiplex relative reverse transcriptase-polymerase chain reaction method. Three of the six genes examined showed modest but consistent upregulation in Se deficiency. Transferrin mRNA was 30% more abundant in Se-deficient than in Se-adequate liver. For the transferrin receptor, the difference was 32%, and for iron regulatory protein 1, it was 63%. No consistent differences were observed for iron regulatory protein 2 or for ferritin light or heavy chain. These findings suggest a possible role for dietary Se in moderating iron metabolism.
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PMID:Selenium regulates expression in rat liver of genes for proteins involved in iron metabolism. 1104

The use of reduced doses of Ritonavir (RIT) and Saquinavir (SQV) is considered a potent alternative in treating patients infected by HIV-1. We tested a combination of 300mg of RIT plus 600mg of SQV, twice daily, in association with two reverse transcriptase inhibitors to treat AIDS patients for a period of 6 months. Evaluation of HIV-1 RNA plasma levels, CD4+/CD8+ cell count and biochemical/hematological parameters (liver enzymes, serum electrolytes, creatinin, blood glucose, uric acid, white blood cell count, platelet count, and hemoglobin level) were performed after 30, 90 and 180 days of therapy. Clinical failure and adverse reactions were also recorded in order to assess safety and efficacy of the treatment. A total of 30 AIDS patients (25 male; 5 female) were enrolled in the study. Eight patients discontinued the therapy due to intolerance, 2 patients presented clinical failure (onset of AIDS-defining events during the study period), 2 patients werc excluded due to protocol violation. Five patients tolerated only a lower dose of RIT (400mg/day). Patients who completed 6 months of therapy had a drop in viral load from 4.8+/-.7 log(10) median 4.9 log) to 3.4 +/- 1.0 log(10) (median 2.6 log), and an increase in CD4+ count from 109 +/- 86cells/ml (median 84cells/ml) to 249+/- 114 cells/ml (median 265 cells/ml), compared to baseline values. However, patients who used a lower dose of RIT (400mg/day) had a less impressive drop in viral load values (mean 0.6 log(10) NA copies/ml) when compared with those using the 600mg/day of the drug (mean 2.4 log(10)). The percentage of patients presenting undetectable levels of HIV-1 RNA in plasma was quite different for the 2 groups: 92% of patients with a viral load <400 RNA copies/ ml were using 600mg of RIT. The combination of reduced doses of RIT and SQV reduced viral load >1.0 log(10) after 6 months in 83% of study patients. The dose of 600mg/day of RIT was morc effective in reducing viral load than 400mg/day, but was less well-tolerated. CD4+ cell counts increased in all patients regardless of the RIT dose used.
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PMID:Safety and Efficacy of Reduced Doses of Ritonavir (RTV) Plus Saquinavir (SQV) in the Treatment of AIDS Patients in Brazil. 1109 12

A 29-year-old woman having acute myelogeneous leukemia-M1 subtype with the chromosomal abnormality t(16;21)(p11;q22) is presented. Complete blood count at onset showed a hemoglobin level of 7.2 g/dl, a platelet count of 48 x 10(9)/l, and a white blood cell count of 161.2 x 10(9)/l with 99% blasts and 1% lymphocytes. Bone marrow aspiration revealed massive proliferation of blasts that were positive for CD13, CD33, CD34, CD56 and myeloperoxidase, and negative for other T-cell, B-cell and monocytic markers. After achieving complete remission following conventional chemotherapy, she received an HLA-matched bone marrow transplantation (BMT) from her sibling after conditioning with busulfan, etoposide and cyclophosphamide. However, 9 months later, the leukemia relapsed as a painful extramedullary mass in her left femur. In spite of intensive re-induction chemotherapy, she died of progressive disease and sepsis. Although we could not detect the TLS/FUS-ERG fusion transcripts by reverse transcriptase-polymerase chain reaction in pre-BMT remission phase, they were clearly detectable in bone marrow cells obtained 6 months after transplantation with no translocation detected by conventional cytogenetics. We consider that even high-dose chemotherapy with BMT may not be effective in the eradication of this type of leukemia, and that the detection of minimal residual disease possibly contributes to the better planning of the therapeutic strategy.
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PMID:Detection of minimal residual disease in a patient having acute myelogenous leukemia with t(16;21)(p11;q22) treated by allogeneic bone marrow transplantation. 1134 Feb 53

Neisseria gonorrhoeae is capable of utilizing host iron-binding proteins, such as transferrin, lactoferrin, and hemoglobin, as the sole source of iron. The receptor involved in transferrin iron acquisition is composed of two distinct transferrin-binding proteins, TbpA and TbpB. The genes that encode these proteins are linked on the chromosome in the order tbpB-tbpA but are separated by an inverted repeat of unknown function. In this study, we sought to understand the transcriptional organization and regulation of the tbp genes, using a combination of lacZ transcriptional fusion analysis and reverse transcriptase PCR (RT-PCR). First, we demonstrated that tbpB and tbpA are cotranscribed and coregulated from the common upstream promoter that precedes tbpB. Using beta-galactosidase activity as a surrogate for tbp-specific transcription, we found that tbpB-specific transcripts were more prevalent than tbpA-specific transcripts after 2 h of growth under iron stress conditions. We confirmed the results obtained by fusion analysis by using RT-PCR applied to native RNA isolated from wild-type gonococci. Three different varieties of RT-PCR were employed: relative, competitive, and real time quantitative. The results of all analyses indicated that tbpB-specific transcripts were approximately twofold more prevalent than tbpA-specific transcripts at steady state. In iron-stressed cultures, the ratio of tbpB- to tbpA-specific message was approximately 2; however, in iron-replete cultures, this ratio dropped to 1. Using these techniques, we also quantitated the effects of iron, external pH, and presence of ligand on tbp mRNA levels.
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PMID:Gonococcal genes encoding transferrin-binding proteins A and B are arranged in a bicistronic operon but are subject to differential expression. 1155 78

Erythrocytes contain a plasma membrane redox system that can reduce extracellular ascorbate radicals by using intracellular ascorbate as an electron donor. In this study, the hypothesis was tested that cytochrome b561 was a component of this system. Spectroscopic analysis of erythrocyte membrane preparations revealed the presence of cytochrome b5 and hemoglobin but also of a cytochrome with properties similar to cytochrome b561, reducible by ascorbate and insensitive to CO. The presence of cytochrome b561 was studied further by reverse transcriptase-PCR analysis of erythrocyte progenitor cells, reticulocytes. However, no cytochrome b561 mRNA could be found. These results were corroborated by Western blot analysis with an anti-cytochrome b561 serum. No cytochrome b561 protein could be detected in extracts of erythrocyte membranes. It is therefore concluded that erythrocytes do not contain cytochrome b561 in their membranes. The possible involvement of other b-cytochromes in ascorbate-ascorbate free radical oxidoreductase activity is discussed.
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PMID:The ascorbate: ascorbate free radical oxidoreductase from the erythrocyte membrane is not cytochrome b561. 1173 44

LuxS is responsible for the production of autoinducer 2 (AI-2), which functions in Vibrio harveyi as a quorum-sensing signal that controls the cell density-dependent expression of the lux operon. In nonluminescent organisms, the physiologic role of AI-2 is not clear. We report that inactivation of luxS in Actinobacillus actinomycetemcomitans JP2 results in reduced growth of the mutant, but not the wild-type organism, under aerobic, iron-limited conditions. Stunted cultures of the luxS mutant A. actinomycetemcomitans JP2-12 grew to high cell density when subcultured under iron-replete conditions. In addition, the mutant strain grew to high cell density under iron limitation after transformation with a plasmid containing a functional copy of luxS. Results of real-time PCR showed that A. actinomycetemcomitans JP2-12 exhibited significantly reduced expression of afuA (eightfold), fecBCDE (10-fold), and ftnAB (>50-fold), which encode a periplasmic ferric transport protein, a putative ferric citrate transporter, and ferritin, respectively. The expressions of putative receptors for transferrin, hemoglobin, and hemophore binding protein were also reduced at more modest levels (two- to threefold). In contrast, expressions of sidD and frpB (encoding putative siderophore receptors) were increased 10- and 3-fold, respectively, in the luxS mutant. To better understand the mechanism of the AI-2 response, the A. actinomycetemcomitans genome was searched for homologs of the V. harveyi signal transduction proteins, LuxP, LuxQ, LuxU, and LuxO. Interestingly, ArcB was found to be most similar to LuxQ sensor/kinase. To determine whether arcB plays a role in the response of A. actinomycetemcomitans to AI-2, an arcB-deficient mutant was constructed. The isogenic arcB mutant grew poorly under anaerobic conditions but grew normally under aerobic iron-replete conditions. However, the arcB mutant failed to grow aerobically under iron limitation, and reverse transcriptase PCR showed that inactivation of arcB resulted in decreased expression of afuA and ftnAB. Thus, isogenic luxS and arcB mutants of A. actinomycetemcomitans exhibit similar phenotypes when cultured aerobically under iron limitation, and both mutants exhibit reduced expression of a common set of genes involved in the transport and storage of iron. These results suggest that LuxS and ArcB may act in concert to control the adaptation of A. actinomycetemcomitans to iron-limiting conditions and its growth under such conditions.
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PMID:luxS and arcB control aerobic growth of Actinobacillus actinomycetemcomitans under iron limitation. 1249 79


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