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)

Dyskeratosis congenita is a rare inherited disorder characterized by abnormal skin manifestations. Morbidity and mortality from this disease is usually due to bone marrow failure, but idiopathic pulmonary fibrosis and an increased cancer predisposition also occur. Families with autosomal dominant dyskeratosis congenita display anticipation and have mutations in the telomerase RNA gene. We identified a three-generation pedigree with autosomal dominant dyskeratosis congenita, anticipation, and telomere shortening. We show that a null mutation in motif D of the reverse transcriptase domain of the protein component of telomerase, hTERT, is associated with this phenotype. This mutation leads to haploinsufficiency of telomerase, and telomere shortening occurs despite the presence of telomerase. This finding emphasizes the importance of telomere maintenance and telomerase dosage for maintaining tissue proliferative capacity and has relevance for understanding mechanisms of age-related changes.
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PMID:Haploinsufficiency of telomerase reverse transcriptase leads to anticipation in autosomal dominant dyskeratosis congenita. 1624 10

Ionizing radiation has been reported to promote accelerated or premature senescence in both normal and tumour cell lines. The current studies were designed to characterize the accelerated senescence response to radiation in the breast tumour cell in terms of its dependence on functional p53 and its relationship to telomerase activity, telomere lengths, expression of human telomerase reverse transcriptase (hTERT, the catalytic subunit of telomerase) and human telomerase RNA (hTR, the RNA subunit of telomerase), as well as the induction of cytogenetic aberrations. Studies were performed in p53 wild-type MCF-7 cells, MCF-7/E6 cells with attenuated p53 function, MDA-MB231 cells with mutant p53 and MCF-7/hTERT cells with constitutive expression of hTERT. Telomerase activity was measured by the telomeric repeat amplification protocol (TRAP assay), telomere lengths by the terminal restriction fragment (TRF) assay, hTR and hTERT expression by reverse transcriptase-polymerase chain reaction (RT-PCR), senescence by beta-galactosidase staining, and apoptosis by TdT-mediated d-UTP-X nick-end labelling (TUNEL assay). Widespread and extensive expression of beta-galactosidase, a marker of cellular senescence, was evident in MCF-7 breast tumour cells following exposure to 10 Gy of ionizing radiation. Radiation did not suppress expression of either hTERT or hTR, alter telomerase activity or induce telomere shortening. Senescence arrest was also observed in irradiated MCF-7/hTERT cells, which have elongated telomeres due to the ectopic expression of the catalytic component of telomerase. In contrast to MCF-7 cells, irradiated MDA-MB231 breast tumour cells and MCF-7/E6 cells failed to senesce and instead demonstrated a delayed apoptotic cell death. Irradiation produced chromosome end associated abnormalities, including end-to-end fusions (an indicator of telomere dysfunction) in MCF-7 cells, MCF-7/hTERT cells, as well as in MCF-7/E6 cells. When cells were maintained in culture following irradiation, proliferative recovery was evident exclusively after senescence while the cell lines which responded to radiation by apoptosis continued to decline in cell number. Accelerated senescence in response to ionizing radiation is p53 dependent and associated with telomer dysfunction but is unrelated to changes in telomerase activity or telomere lengths, expression of hTERT and hTR. In the absence of functional p53, cells are unable to arrest for an extended period, resulting in apoptotic cell death while accelerated senescence in cells expressing p53 is succeeded by proliferative recovery.
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PMID:p53-Dependent accelerated senescence induced by ionizing radiation in breast tumour cells. 1630 15

Telomerase is essential for immortalization of most human cancer cells. Expression of the core telomerase RNA (hTR) and reverse transcriptase (hTERT) subunits is mainly regulated by transcription. However, hTR transcriptional regulation remains poorly understood. We previously showed that the core hTR promoter is activated by Sp1 and is repressed by Sp3. Here, we show that the mitogen-activated protein kinase kinase kinase 1 (MEKK1)/c-Jun-NH(2)-kinase (JNK) pathway represses hTR expression by a mechanism that involves Sp1 and Sp3. Promoter activity was induced by the JNK inhibitor SP600125 and was repressed by activated MEKK1. Repression by MEKK1 was blocked by SP600125 or enhanced by coexpression of wild-type but not phosphoacceptor mutated JNK. SP600125 treatment also increased levels of endogenous hTR. Mutations in the hTR promoter Sp1/Sp3 binding sites attenuated SP600125-mediated promoter induction, whereas coexpression of MEKK1 with Sp3 enhanced hTR promoter repression. Chromatin immunoprecipitation showed that levels of immunoreactive Sp1 associated with the hTR promoter were low in comparison with Sp3 in control cells but increased after JNK inhibition with a reciprocal decrease in Sp3 levels. No corresponding changes in Sp1/Sp3 protein levels were detected. Thus, JNK represses hTR promoter activity and expression, apparently by enhancing repression through Sp3.
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PMID:Transcriptional repression of telomerase RNA gene expression by c-Jun-NH2-kinase and Sp1/Sp3. 1645 90

A number of previous studies investigated the in vitro effects of resveratrol on malignant human breast epithelial cell replication. The aim of the present study was to evaluate the activity of resveratrol on human metastatic breast cancer cells. The study was performed on the MCF-7 tumor cell line. Cell growth, cell cycle perturbation and apoptosis were evaluated by trypan blue dye exclusion assay, flow cytometric analysis and confocal fluorescence microscopy. TRAP assay and Western blot analysis respectively detected levels of telomerase activity and levels of hTERT in intracellular compartments of MCF-7 cells treated with resveratrol. Resveratrol has a direct inhibitory effect on cell proliferation. The results demonstrate that the drug induces apoptosis in MCF-7 cells, in a time- and concentration-related manner. Our results also show that the growth-inhibitory effect of resveratrol on malignant cells is mainly due to its ability to induce S-phase arrest and apoptosis in association with reduced levels of telomerase activity. In particular, TRAP assay and Western blot analysis respectively showed that resveratrol treatment down-regulates the telomerase activity of target cells and the nuclear levels of hTERT, the reverse transcriptase subunit of the telomerase complex. In our experimental model of breast cancer, resveratrol shows direct antiproliferative and pro-apoptotic effects. Studies on telomerase function and intracellular hTERT distribution point out that this agent is endowed with additional suppressive functions on critical tumor biological properties. These results speak in favor of a potential role of resveratrol in chemoprevention/chemotherapy of breast cancer.
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PMID:Resveratrol down-regulates the growth and telomerase activity of breast cancer cells in vitro. 1646 68

One of the challenges of oncolytic virotherapy is the inability to easily track or monitor virus activity during treatment. Here we describe the construction and functional characterization of Ad/hTC-GFP-E1, an oncolytic virus whose transgenes GFP and E1A are both under the control of a synthetic promoter (hTC). This promoter consists of sequences from the human telomorase reverse transcriptase promoter and a minimal cytomegalovirus (CMV) early promoter. The tumor-specific expression of E1A and GFP was demonstrated by Western blot and fluorescent microscope analyses, and the tumor-specific cytotoxicity by crystal-violet staining and cell viability assays. Viral replication and tumor cell lysis occurred at multiplicities of infection (MOI) as low as 100 viral particles per cell in sensitive cell lines. No overt cytotoxic effect was observed in normal human fibroblasts, even at MOIs over 2000 vp. The presence of oncolytic vector was easily visualized and quantitated in vitro and in vivo, in correlation with viral replication. Intralesional administration of the virus into subcutaneous H1299 (NSCLC) tumor xenografts significantly suppressed tumor growth and provided a survival benefit. Together, these results demonstrate that an hTERT-specific oncolytic adenovirus expressing an hTERT-specific transgene is applicable for cancer therapy.
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PMID:Oncolysis and suppression of tumor growth by a GFP-expressing oncolytic adenovirus controlled by an hTERT and CMV hybrid promoter. 1648 10

Genistein, the most abundant isoflavone present in soybean has antiproliferative effects on a variety of cancer cells, including prostate cancer. However, the molecular mechanism of antiproliferative effects of genistein is not entirely understood. Because the activation of telomerase is crucial for cells to gain immortality and proliferation ability, we examined the role of genistein in the regulation of telomerase activity in prostate cancer cells. Here, we show that genistein-induced inhibition in cell proliferation is associated with a reduction in telomerase activity. Using reverse transcriptase-PCR and hTERT promoter activity assays, we showed that genistein decreased hTERT expression and transcriptional activity dose-dependently. Using various deleted hTERT promoter constructs, we defined that the hTERT core promoter is enough to observe the genistein-induced repression of hTERT transcriptional activity. Because c-Myc is involved in transcriptional regulation of hTERT, c-Myc expression was examined. A dose-dependent decrease in c-Myc message and proteins was observed with genistein treatment. These results indicate that genistein represses hTERT transcriptional activity via the down-regulation of c-Myc expression. However, genistein-induced repression of hTERT transcriptional activity was not blocked by the mutation of c-Myc at the hTERT promoter, suggesting that additional factors are involved in genistein-dependent repression of telomerase activity. Interestingly, we observed that genistein down-regulates the activation of Akt thereby phosphorylation of hTERT and inhibits its translocation to the nucleus. These results show for the first time that genistein represses telomerase activity in prostate cancer cells not only by repressing hTERT transcriptional activity via c-Myc but also by posttranslational modification of hTERT via Akt.
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PMID:Genistein represses telomerase activity via both transcriptional and posttranslational mechanisms in human prostate cancer cells. 1648 11

Telomeres are located at the ends of eukaryotic chromosomes. Human telomerase, a cellular reverse transcriptase, is a ribonucleoprotein enzyme that catalyzes the synthesis and extension of telomeric DNA. It is composed of at least, a template RNA component (hTR; human Telomerase RNA) and a catalytic subunit, the telomerase reverse transcriptase (hTERT). The absence of telomerase is associated with telomere shortening and aging of somatic cells, while high telomerase activity is observed in over 85% of human cancer cells, strongly indicating its key role during tumorigenesis. Several details regarding telomere structure and telomerase regulation have already been elucidated, providing new targets for therapeutic exploitation. Further support for anti-telomerase approaches comes from recent studies indicating that telomerase is endowed of additional functions in the control of growth and survival of tumor cells that do not depend only on the ability of this enzyme to maintain telomere length. This observation suggests that inhibiting telomerase or its synthesis may have additional anti-proliferative and apoptosis inducing effect, independently of the reduction of telomere length during cell divisions. This article reviews the basic information about the biology of telomeres and telomerase and attempts to present various approaches that are currently under investigation to inhibit its expression and its activity. We summarize herein distinct anti-telomerase approaches like antisense strategies, reverse transcriptase inhibitors, and G-quadruplex interacting agents, and also review molecules targeting hTERT expression, such as retinoids and evaluate them for their therapeutic potential. "They conceive a certain theory, and everything has to fit into that theory. If one little fact will not fit it, they throw it aside. But it is always the facts that will not fit in that are significant". "Death on the Nile". Agatha Christie.
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PMID:Telomeres and telomerase: Pharmacological targets for new anticancer strategies? 1652 44

Telomeres are repetitive DNA sequences at the ends of linear chromosomes. Telomerase, a cellular reverse transcriptase, helps maintain telomere length in human stem cells, reproductive cells and cancer cells by adding TTAGGG repeats onto the telomeres. However, most normal human cells do not express telomerase and thus each time a cell divides some telomeric sequences are lost. When telomeres in a subset of cells become short (unprotected), cells enter an irreversible growth arrest state called replicative senescence. Cells in senescence produce a different constellation of proteins compared to normal quiescent cells. This may lead to a change in the homeostatic environment in a tissue-specific manner. In most instances cells become senescent before they can become cancerous; thus, the initial growth arrest induced by short telomeres may be thought of as a potent anti-cancer protection mechanism. When cells can be adequately cultured until they reach telomere-based replicative senescence, introduction of the telomerase catalytic protein component (hTERT) into telomerase-silent cells is sufficient to restore telomerase activity and extend cellular lifespan. Cells with introduced telomerase are not cancer cells, since they have not accumulated the other changes needed to become cancerous. This indicates that telomerase-induced telomere length manipulations may have utility for tissue engineering and for dissecting the molecular mechanisms underlying genetic diseases, including cancer.
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PMID:Hallmarks of telomeres in ageing research. 1720 Sep 48

The telomerase RNA (hTR) and reverse transcriptase (hTERT) promoters are active in most cancer cells, but not in normal cells, and are useful for transcriptional targeting in gene therapy models. Telomerase-specific conditionally replicating adenoviruses (CRAd) are attractive vectors because they should selectively lyse tumor cells. Here, we compare CRAds, in which either the hTR or hTERT promoter controls expression of the adenovirus E1A gene. In replication-defective reporter adenoviruses, the hTR promoter was up to 57-fold stronger in cancer cells than normal cells and up to 49-fold stronger than hTERT. In normal cells, hTERT promoter activity was essentially absent. Doses of telomerase-specific CRAds between 1.8 and 28 infectious units per cell efficiently killed cancer cells, but normal cells required higher doses. However, CRAd DNA replication and E1A expression were detected in both cancer and normal cells. Overall, tumor specificity of the CRAds was limited compared with nonreplicating vectors. Surprisingly, both CRAds expressed similar E1A levels and functional behavior, despite known differentials between hTR and hTERT promoter activities, suggesting that the promoters are deregulated. Rapid amplification of cDNA ends analysis of hTR-/hTERT-E1A transcripts ruled out cryptic transcription from the vector backbone. Blocking E1A translation partially restored the hTR-/hTERT-E1A mRNA differential, evidencing feedback regulation by E1A.
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PMID:Modulation of telomerase promoter tumor selectivity in the context of oncolytic adenoviruses. 1728 67

Normal human somatic cells undergo limited cell division cycles and enter irreversible replication arrest called senescence. Cellular senescence of many human cell types is regulated by the length and status of telomeric sequences, which is shortened after each round of DNA replication. Telomeres can be rejuvenated by telomerase, an enzyme which carries out de novo synthesis of telomeric DNA. Telomerase is a ribonucleoprotein complex composed minimally of telomere reverse transcriptase gene (hTERT) and RNA template (hTR), and its enzyme activity in cells is primarily limited by the level of hTERT expression. Therefore, telomerase activity in cells can be reconstituted by overexpression of hTERT, frequently resulting in extension of replicative life span or immortalization. It is well established that the effect of telomerase reconstitution on cellular life span is clearly cell type-dependent because telomere shortening is not the only limiting factor of cellular life span. However, telomerase activity appears to be a requirement for cellular immortalization, irrespective of the cell types. In this article, we discuss the detailed methods to extend the in vitro replicative life span of primary human cells by ectopic expression of hTERT.
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PMID:Extension of cell life span using exogenous telomerase. 1763 80


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