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)

Telomeres are specialized structures at the end of eukaryotic chromosomes that in vertebrates constain hundreds to thousands of tandem repeats of the sequence TTAGGG. In most human somatic cells, telomeres shorten with each cell division, eventually triggering an irreversible arrest of proliferation called cellular senescence. These observations have led to a model in which telomere length reflects the mitotic history of somatic cells. Further support for this hypothesis has come from the discovery of telomerase, a unique reverse transcriptase ribonucleoprotein that has the ability to extend 3' end of telomeres. In fibroblasts, senescence is induced by telomere attrition and depends on p53 and pRb pathways triggered by one or a few critically short telomeres. Previous studies have shown that the replicative life span of various primary human cells can be prolonged by transduction of the telomerase reverse transcriptase (hTERT) gene. The hTERT expressing cells proliferate indefinitely, without undergoing any changes associated with transformation to malignancy. Rapid progress has been made towards the goal of using tumor-specific cytolytic CD8+ T lymphocytes for the immunotherapy of cancer. These cells can be expanded in vitro and, in principle, could be used for adoptive immunotherapy. One of the major problems that remains to be solved is the finite life span of normal human T lymphocytes. In an attempt to overcome this barrier three groups have introduced hTERT cDNA into human T lymphocytes and monitored its effect on their life span. In two of these studies, hTERT significantly extended the replicative life span of CD8+ T clones, whereas this was not the case in the third study using bulk T lymphocytes. Possible explanations for these discordant results are that better growth conditions avoided culture-induced stress in the study with clones, or that clones had undergone alterations leading, for example, to the inactivation of the pRb pathway during their derivation.
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PMID:[Telomerase, elixir of life for human cells?]. 1283 17

Telomerase is an RNA-dependent DNA polymerase that synthesizes TTAGGG telomeric DNA onto chromosome ends to compensate for sequence loss during DNA replication. It has been detected in 85-90% of all primary human cancers, implicating that the telomerase seems to be reactivated in tumors and that such activity may play a role in the tumorigenic process. The purpose of this study was to evaluate telomerase activity, human telomerase RNA (hTR), and telomerase reverse transcriptase (TERT) in stomach cancer and to determine their potential relationships to clinicopathologic parameters. Frozen and corresponding methacarn-fixed paraffin-embedded tissue samples were obtained from 51 patients with gastric adenocarcinoma and analyzed for telomerase activity by using a TRAPeze ELISA kit. Tissue sections of all the samples were further investigated for hTR and TERT by in situ hybridization and a sensitive immunohistochemical technique, respectively. Telomerase activity was detected in 37 (73%) tumors. Telomerase positivity from methacarn-fixed paraffin blocks was found to be 35% of that from frozen tissues. hTR was overexpressed in 46 (90%) samples: 33/37 (89%) with and 13/14 (93%) without telomerase activation. Expression of TERT was demonstrated in 40 (78%) cases: 30/37 (81%) with and 10/14 (71%) without telomerase. Telomerase activity correlated well with depth of invasion (P =.037) and tumor differentiation (P =.022), whereas hTR significantly correlated with nodal metastasis (P=.047) and tumor size (P=.023). These data suggest that reactivated telomerase may play a significant role in the tumorigenesis of gastric cancer and may reflect, along with enhanced hTR, the malignant potential of the tumor. It is noteworthy that methacarn-fixed tissue cannot as yet substitute for the frozen section in the TRAP assay.
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PMID:Expression of telomerase activity, human telomerase RNA, and telomerase reverse transcriptase in gastric adenocarcinomas. 1286 Oct 67

Telomerase is an enzyme that utilizes an internal RNA molecule as a template for the extension of chromosomal DNA ends. The catalytic core of telomerase consists of the RNA subunit and a protein reverse transcriptase subunit, known as telomerase reverse transcriptase (TERT). It has previously been shown that both yeast and human telomerase can form dimers or multimers in which one RNA in the complex can influence the activity of another. To test the proposal that dimerization might be essential for telomerase activity, we sought to determine whether Tetrahymena thermophila telomerase is active as a dimer or a monomer. Recombinant Tetrahymena telomerase eluted from a gel filtration column at the size of a monomeric complex (one RNA plus one TERT), and those fractions showed processive telomerase activity. We were unable to detect dimerization of Tetrahymena telomerase by coprecipitation experiments, by using tags on either the TERT protein or telomerase RNA. Therefore, a majority, if not all, of the recombinant Tetrahymena telomerase in our reconstitution system is present as a monomeric complex. We were also unable to detect dimerization of native telomerase from mating and vegetative Tetrahymena cell extracts. These results demonstrate that Tetrahymena telomerase does not need to dimerize to be active and processive.
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PMID:Tetrahymena telomerase is active as a monomer. 1367 9

The reverse transcriptase telomerase copies an internal RNA template to synthesize telomeric simple-sequence repeats. In the cellular context, telomerase must elongate its few intended substrates (authentic chromosome ends) without spurious activity on other potential substrates (chromosome ends created by damage, repair, or recombination). Many mechanisms have been proposed to account for the biological substrate specificity of telomerase, with most models focusing on protein-protein interactions between telomerase and telomeric chromatin. Telomerase activity assays testing the elongation of model oligonucleotide substrates have revealed that in addition to hybridization with the RNA template, optimal DNA substrates also engage telomerase protein-based interaction sites. The physiological significance of these non-template interaction sites has not been established. We used in vivo reconstitution to assemble telomerase enzymes with variant telomerase reverse transcriptase proteins. Several telomerase enzyme variants retained a wild-type level of catalytic function in vitro when assayed using an artificial sequence substrate but exhibited reduced activity on a more physiological telomeric-sequence substrate. Telomerases that demonstrated this defect in telomeric substrate usage in vitro also failed to support telomere length maintenance in vivo. Our findings suggest that non-template interactions of the telomerase ribonucleoprotein with telomeric DNA play a critical role in supporting telomerase function on its appropriate cellular substrates.
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PMID:Human telomerase reverse transcriptase motifs required for elongation of a telomeric substrate. 1456 61

Telomerase is a ribonuclear protein reverse transcriptase that maintains telomere length in eukaryotic cells. Activation of telomerase has been implicated in human cellular immortalization and carcinogenesis. Telomerase activity in ovarian neoplasm has been studied using polymerase chain reaction (PCR)-based methods and shown to be correlated with malignancy. However, we believe those results must be interpreted with caution because such studies used a heterogeneous mix of cells, including normal cell type known to express telomerase when activated. The present study used in situ hybridization that allows determination of the type of cells expressing telomerase, as well as the intensity of that expression, in ovarian neoplasms. A total of 75 specimens were studied. Epithelial telomerase reverse transcriptase mRNA expression was detected in 28 of 31 epithelial ovarian carcinomas, 1 of 1 malignant granulosa cell tumor, 7 of 9 serous borderline ovarian tumors, 11 of 11 mucinous borderline ovarian tumors, 4 of 5 serous cystadenofibromas, 2 of 4 serous cystadenomas, 8 of 8 mucinous cystadenomas, and 0 of 6 normal ovaries except the corpus luteum. Telomerase expression is heterogeneously found in both benign and malignant epithelial tissues. We conclude that human telomerase reverse transcriptase mRNA expression does not seem to be a reliable marker for clinical use in differentiating between benign and malignant tumors.
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PMID:In situ human telomerase reverse transcriptase expression pattern in normal and neoplastic ovarian tissues. 1471 58

BACKGROUND: Telomerase is a ribonucleoprotein enzyme that synthesises telomeres after cell division and maintains chromosomal stability leading to cellular immortalisation. hTERT (human telomerase reverse transcriptase) is the rate-limiting determinant of telomerase reactivation. Telomerase has been associated with negative prognostic indicators in some studies. The present study aims to detect any correlation between hTERT and the negative prognostic indicators VEGF and PCNA by quantitatively measuring the mRNA expression of these genes in human breast cancer and in adjacent non-cancerous tissue (ANCT). MATERIALS AND METHODS: RNA was extracted from 38 breast carcinomas and 40 ANCT. hTERT and VEGF165, VEGF189 and PCNA mRNA expressions were estimated by reverse transcriptase-PCR (RT-PCR) and Taqman methodology. RESULTS: The level of expression of VEGF-165 and PCNA was significantly higher in carcinoma tissue than ANCT (p = 0.02). The ratio of VEGF165/189 expression was significantly higher in breast carcinoma than ANCT (p = 0.025). hTERT mRNA expression correlated with VEGF-189 mRNA (p = 0.008) and VEGF165 (p = 0.07). CONCLUSIONS: hTERT mRNA expression is associated with the expression of the VEGF189 and 165 isoforms. This could explain the poorer prognosis reported in breast tumours expressing high levels of hTERT. The relative expression of the VEGF isoforms is significantly different in breast tumour to ANCT, and this may be important in breast carcinogenesis.
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PMID:The mRNA expression of hTERT in human breast carcinomas correlates with VEGF expression. 1473 67

Primitive neuroectodermal tumors (PNETs), including medulloblastoma (PNET/MB) and supratentorial PNET (sPNET), are the most common malignant brain tumors of childhood. The stabilization of telomere lengths by telomerase activation is an important step in carcinogenesis and cell immortalization. Epigallocatechin gallate (EGCG), the major polyphenol in green tea, is a telomerase inhibitor with antiproliferative and anticarcinogenic effects against different types of cancer. In this study, we used real-time reverse transcriptase-polymerase chain reaction to measure the mRNA expression of the human telomerase reverse transcriptase (hTERT) in 50 primary PNET samples (43 PNET/MB, 7 sPNET), 14 normal human brain samples, and 6 human PNET cell lines. Compared to normal human cerebellum, 38/50 (76%) primary PNET samples had >or= 5-fold upregulated hTERT mRNA expression. We then examined PNET cell lines for telomerase activity using a quantitative telomeric repeat amplification protocol (TRAP), and for telomere length using terminal restriction fragment analysis. While a positive correlation between hTERT mRNA expression and telomerase activity was detected in PNET cell lines, no correlation was found between telomerase activity and telomere length. Treatment of PNET cell lines with EGCG resulted in a dose-dependent inhibition of telomerase activity at micromolar levels. Although EGCG displayed strong proliferation inhibitory effects against TRAP-positive PNET cell lines, it had no significant effect against TRAP-negative D425 cells. These results provide evidence for a possible role of telomerase in the pathogenesis of most PNETs and indicate that subsets of PNETs maintain telomere length by alternative mechanisms. Inhibition of telomerase function represents a novel experimental therapeutic strategy in childhood PNETs that warrants further investigation.
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PMID:Telomere maintenance in childhood primitive neuroectodermal brain tumors. 1476 33

There is accumulating evidence that peptides derived from the catalytic subunit of human telomerase reverse transcriptase (hTERT) are specifically recognized by CD8+ cytotoxic T lymphocytes. We investigated the cytotoxicity of a human leukocyte antigen (HLA)-A*2402-restricted hTERT-derived peptide 461-469 (hTERT461)-specific CD8+ T-cell clone, designated as K3-1, established from a healthy donor by repetitive peptide stimulation. This clone exhibited cytotoxicity against 4 out of 6 HLA-A24-positive lung cancer cell lines with positive telomerase activity but not 4 HLA-A24-negative examples. When the target cells were pretreated with 100 U/ml of interferon (IFN)-gamma for 48 hr, the susceptibility to K3-1 increased with PC9 cells but unexpectedly decreased with LU99 cells. However, in both cell lines, the expression of molecules associated with epitope presentation such as HLA-A24, transporters associated with antigen processing, low molecular weight polypeptide 7 and proteasome activator 28 was similarly increased after IFN-gamma treatment. Results of CTL assays using acid-extracted peptides indicated that the epitope increased on PC9 cells but not on LU99 cells after IFN-gamma treatment. Semi-quantitative reverse transcriptase polymerase chain reaction disclosed that the expression of hTERT was attenuated in LU99 but not in PC9 cells, accounting for the decreased cytotoxicity mediated by K3-1. The attenuation of the hTERT expression and K3-1-mediated cell lysis after IFN-gamma treatment was also observed in primary adenocarcinoma cells obtained from pulmonary fluid of a lung cancer patient. Our data underline the utility of peptide hTERT461 in immunotherapy for lung cancer, as with other malignancies reported earlier, and suggest that modulation of hTERT expression by IFN-gamma needs to be taken into account in therapeutic approach.
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PMID:Interferon-gamma differentially regulates susceptibility of lung cancer cells to telomerase-specific cytotoxic T lymphocytes. 1509 6

Telomerase is a ribonucleoprotein complex mainly composed of a reverse transcriptase catalytic subunit (telomerase reverse transcriptase gene, hTERT) that copies a template region of its RNA subunit to the end of the telomere. For detecting telomerase activity in a tissue specimen the TRAP assay is a relatively sensitive and specific method, but it can be used only on fresh tissue extracts and offers no information at the single cell level. Immunohistochemistry (IHC) allows to detect hTERT protein expression at an individual cell level in human tissues. We have tested commercially available anti-hTERT antibodies in formalin-fixed and paraffin-embedded human tissues by IHC. Only one monoclonal antibody (NCL-hTERT; Novacastra) was sufficiently specific and this was applied to human tissues in which telomerase activity had been shown by TRAP assay and hTERT mRNA expression by RT-PCR. hTERT protein localized diffusely in the nucleoplasm and more intensely in the nucleoli of cancer cells and proliferating normal cells. Mitotic cells showed diffuse staining of the entire cell. Granular cytoplasmic staining was occasionally found in some tumor cells. In telomerase-positive tumors not all the tumor cells showed hTERT immunoreactivity. A significantly heterogeneous hTERT protein expression was observed in human tumor tissues. The hTERT immunostaining in fixed tissues was concordant with telomerase activity and hTERT mRNA expression in corresponding non-fixed samples. Quantitative RT-PCR of microdissected sections showed that hTERT mRNA expression was higher in cells with nuclear expression than in those with cytoplasmic expression. Double staining with the M30 antibody showed that a subpopulation of hTERT-negative cells is apoptotic. We conclude that: (1) hTERT protein can be detected by IHC in fixed human tissues, but the choice of the antibody, tissue processing, and reaction conditions are critical, (2) hTERT protein localizes in the nucleoplasm, more strongly in the nucleolus, and occasionally in the cytoplasm, (3) telomerase-positive tumors show significant heterogeneity of hTERT protein expression, and (4) a subpopulation of hTERT protein negative tumor cells is identified as apoptotic cells.
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PMID:Immunohistochemical localization of hTERT protein in human tissues. 1513 42

In immortal cells, the existence of a mechanism for the maintenance of telomere length is critical. In most cases this is achieved by the reactivation of telomerase, a cellular reverse transcriptase that prevents telomere shortening. Here we report that the telomerase gene (hTERT) promoter is up-regulated during transmission of human T-cell lymphotropic virus type-I (HTLV-I) to primary T cells in vitro and in ex vivo adult T-cell leukemia/lymphoma (ATLL) samples, but not asymptomatic carriers. Although Tax impaired induction of human telomerase reverse transcriptase (hTERT) mRNA in response to mitogenic stimulation, transduction of Tax into primary lymphocytes was sufficient to activate and maintain telomerase expression and telomere length when cultured in the absence of any exogenous stimulation. Transient transfection assays revealed that Tax stimulates the hTERT promoter through the nuclear factor kappaB (NF-kappaB) pathway. Consistently, Tax mutants inactive for NF-kappaB activation could not activate the hTERT or sustain telomere length in transduced primary lymphocytes. Analysis of the hTERT promoter occupancy in vivo using chromatin immunoprecipitation assays suggested that an increased binding of c-Myc and Sp1 is involved in the NF-kappaB-mediated activation of the hTERT promoter. This study establishes the role of Tax in regulation of telomerase expression, which may cooperate with other functions of Tax to promote HTLV-I-associated adult T-cell leukemia.
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PMID:Transcriptional activation of hTERT through the NF-kappaB pathway in HTLV-I-transformed cells. 1522 82


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