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)

Here, we show that inhibition of c-Myc causes a proliferative arrest of M14 melanoma cells through cellular crisis, evident by the increase in size, multiple nuclei, vacuolated cytoplasm, induction of senescence-associated beta-galactosidase activity and massive apoptosis. The c-Myc-induced crisis is associated with decreased human telomerase reverse transcriptase expression, telomerase activity, progressive telomere shortening, glutathione (GSH), depletion and, increased production of reactive oxygen species. Treatment of control cells with L-buthionine sulfoximine decreases GSH to levels of c-Myc low expressing cells, but it does not modify the growth kinetic of the cells. Surprisingly, when GSH is increased in the c-Myc low expressing cells by treatment with N-acetyl-L-cysteine, cells escape crisis. To test the hypothesis that both oxidative stress and telomerase dysfunction are involved in the c-Myc-dependent crisis, we directly inhibited telomerase function and glutathione levels. Inactivation of telomerase, by expression of a catalytically inactive, dominant negative form of reverse transcriptase, reduces cellular lifespan by inducing telomere shortening. Treatment of cells with L-buthionine sulfoximine decreases GSH content and accelerates cell crisis. Analysis of telomere status demonstrated that oxidative stress affects c-Myc-induced crisis by increasing telomere dysfunction. Our results demonstrate that inhibition of c-Myc oncoprotein induces cellular crisis through cooperation between telomerase dysfunction and oxidative stress.
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PMID:Inhibition of c-Myc oncoprotein limits the growth of human melanoma cells by inducing cellular crisis. 1282 59

The 5' untranslated region of the proto-oncogene c-myc contains an internal ribosome entry segment and c-Myc translation can be initiated by cap-independent as well as cap-dependent mechanisms. In contrast to the process of cap-dependent initiation, the trans-acting factor requirements for cellular internal ribosome entry are poorly understood. Here, we show that members of the poly (rC) binding protein family, poly (rC) binding protein 1 (PCBP1), poly (rC) binding protein 2 (PCBP2) and hnRNPK were able to activate the IRES in vitro up to threefold when added in combination with upstream of N-ras and unr-interacting protein. The interactions of PCBP1, PCBP2 and hnRNPK with c-myc-IRES-RNA were shown to be specific by ultraviolet crosslinking analysis and electrophoretic mobility shift assays, while immunoprecipitation of the three proteins using specific antibodies followed by reverse transcriptase-polymerase chain reaction showed that they were able to bind c-myc mRNA. c-myc-IRES-mediated translation from the reporter vector was stimulated by cotransfection of plasmids encoding PCBP1, PCBP2 and hnRNPK. Interestingly, the mutated version of the c-myc IRES that is prevalent in patients with multiple myeloma bound hnRNPK more efficiently in vitro and was stimulated by hnRNPK to a greater extent in vivo.
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PMID:Members of the poly (rC) binding protein family stimulate the activity of the c-myc internal ribosome entry segment in vitro and in vivo. 1297 Jul 49

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

BACKGROUND: Telomerase is a ribonucleoprotein enzyme that synthesises telomeres after cell division and maintains chromosomal length and stability thus leading to cellular immortalisation. The hTERT (human telomerase reverse transcriptase) subunit seems to be the rate-limiting determinant of telomerase and knowledge of factors controlling hTERT transcription may be useful in therapeutic strategies. The hTERT promoter contains binding sites for c-Myc and there is experimental and in vitro evidence that c-Myc may increase hTERT expression. MATERIALS AND METHODS: RNA was extracted from 18 breast carcinomas and c-Myc mRNA expression was estimated by quantitative reverse transcriptase-PCR (RT-PCR) with Taqman methodology. These tumours had already been analysed for ER and PgR status using ligand-binding assays and had had their DNA ploidy and S-phase fractions measured by flow cytometry. Telomerase activity had already been determined by using a modified telomeric repeat and amplification protocol (TRAP) assay. RESULTS: Telomerase activity ranged from 0 to 246 units of Total Protein Generated (TPG), where one unit of TPG was equal to 600 molecules of telomerase substrate primers extended by at least three telomeric repeats. Median levels of TPG were 60 and mean levels 81. There was no significant correlation between levels of c-Myc mRNA expression, telomerase activity, S phase fraction or PgR. There was a significant negative correlation with ER status. CONCLUSION: Although the hTERT promoter contains potential binding sites for c-Myc oncoprotein, we have found no correlation between c-Myc mRNA levels and telomerase activity.
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PMID:There is no correlation between c-Myc mRNA expression and telomerase activity in human breast cancer. 1528 10

Prostate cancer is the leading cause of cancer-related deaths in men. Androgen ablation is the mainstay of treatment for advanced prostate cancer. This therapy is very effective in androgen-dependent cancer; however, these cancers eventually become androgen independent, rendering anti-androgen therapy ineffective. The exploration of novel modalities of treatment is therefore essential to improve the prognosis of this neoplasia. Telomeres are specialized heterochromatin structures that act as protective caps at the ends of chromosomes. Telomere maintenance in the majority of tumor cells is achieved by telomerase, a reverse transcriptase enzyme that catalyzes the synthesis of further telomeric DNA. Telomerase is detected in the majority of prostate cancers, but not in normal or benign prostatic hyperplasia tissue. Moreover, the human telomerase reverse transcriptase (hTERT) gene, the catalytic subunit of telomerase, is regulated by androgens as well as by different oncogenes including Her-2, Ras, c-Myc and Bcl-2, which seem to play an important role in prostate cancer progression. Thus, telomerase may represent a very good candidate for targeted therapy in prostate tumors. To inhibit telomere maintenance by telomerase, approaches that directly target either telomerase and telomeres or the telomerase regulatory mechanisms have been used. Moreover, strategies targeting telomerase-positive cells as a means to directly kill the tumor cells have been tested. This review summarizes the most promising results achieved by anti-telomerase strategy in different solid tumors. Most of the telomerase-associated therapies described here have proved very promising for the treatment of prostate cancer. On the basis of the good results obtained and considering the multigenic defects of human tumors, including prostate cancer, the combination of anti-telomerase strategies with conventional drugs and/or molecules capable of interfering with oncogenic pathways could efficiently improve the response of this neoplasia.
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PMID:Telomerase as a new target for the treatment of hormone-refractory prostate cancer. 1536 45

Telomerase, the enzyme which maintains the ends of linear chromosomes in eukaryotic cells is found in murine embryonic stem cells; however, its activity is downregulated during in vitro differentiation. Previous work has indicated that this is due to the transcriptional downregulation of murine reverse transcriptase unit (mTert) of telomerase. To investigate the factors that cause the transcriptional repression of mTert we defined a 300 bp region which is essential for its transcription and performed site directed mutagenesis and electrophoretic mobility shift assays. This analysis indicated that Sp1, Sp3 and c-Myc bind to the GC-boxes and E-boxes, respectively, within the promoter and help activate the transcription of mTert gene. We also identified a novel binding sequence, found repeated within the mTert core region, which when mutated caused increased mTert expression. Yeast one hybrid screening combined with electrophoretic mobility shift assays indicated that the nuclear protein Zap3 binds to this site and its overexpression leads to the downregulation of mTert during differentiation. This suggests that regulation of mTert transcription is a complex process which depends on a quantitative balance between transcription factors that cause activation or repression of this gene. Overexpression of Zap3 in murine embryonic stem cells results in reduction in telomerase activity and telomere length as well as reduced proliferative capacity and limited ability to contribute to the development of haematopoietic cells upon differentiation.
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PMID:A role for nucleoprotein Zap3 in the reduction of telomerase activity during embryonic stem cell differentiation. 1551 42

Kaposi's sarcoma-associated herpesvirus (KSHV) in vitro target cell infection is characterized by the expression of the latency-associated genes ORF 73 (LANA-1), ORF 72, and K13 and by the transient expression of a very limited number of lytic genes such as lytic cycle switch gene ORF 50 (RTA) and the immediate early (IE) lytic K5, K8, and v-IRF2 genes. During the early stages of infection, several overlapping multistep complex events precede the initiation of viral gene expression. KSHV envelope glycoprotein gB induces the FAK-Src-PI3K-RhoGTPase (where FAK is focal adhesion kinase) signaling pathway. As early as 5 min postinfection (p.i.), KSHV induced the extracellular signal-regulated kinase 1 and 2 (ERK1/2) via the PI3K-PKCzeta-MEK pathway. In addition, KSHV modulated the transcription of several host genes of primary human dermal microvascular endothelial cells (HMVEC-d) and fibroblast (HFF) cells by 2 h and 4 h p.i. Neutralization of virus entry and infection by PI-3K and other cellular tyrosine kinase inhibitors suggested a critical role for signaling molecules in KSHV infection of target cells. Here we investigated the induction of ERK1/2 by KSHV and KSHV envelope glycoproteins gB and gpK8.1A and the role of induced ERK in viral and host gene expression. Early during infection, significant ERK1/2 induction was observed even with low multiplicity of infection of live and UV-inactivated KSHV in serum-starved cells as well as in the presence of serum. Entry of UV-inactivated virus and the absence of viral gene expression suggested that ERK1/2 induction is mediated by the initial signal cascade induced by KSHV binding and entry. Purified soluble gpK8.1A induced the MEK1/2 dependent ERK1/2 but not ERK5 and p38 mitogen-activated protein kinase (MAPK) in HMVEC-d and HFF. Moderate ERK induction with soluble gB was seen only in HMVEC-d. Preincubation of gpK8.1A with heparin or anti-gpK8.1A antibodies inhibited the ERK induction. U0126, a selective inhibitor for MEK/ERK blocked the gpK8.1A- and KSHV-induced ERK activation. ERK1/2 inhibition did not block viral DNA internalization and had no significant effect on nuclear delivery of KSHV DNA during de novo infection. Analyses of viral gene expression by quantitative real-time reverse transcriptase PCR revealed that pretreatment of cells with U0126 for 1 h and during the 2-h infection with KSHV significantly inhibited the expression of ORF 73, ORF 50 (RTA), and the IE-K8 and v-IRF2 genes. However, the expression of lytic IE-K5 gene was not affected significantly. Expression of ORF 73 in BCBL-1 cells was also significantly inhibited by preincubation with U0126. Inhibition of ERK1/2 also inhibited the transcription of some of the vital host genes such as DUSP5 (dual specificity phosphatase 5), ICAM-1 (intercellular adhesion molecule 1), heparin binding epidermal growth factor, and vascular endothelial growth factor that were up-regulated early during KSHV infection. Several MAPK-regulated host transcription factors such as c-Jun, STAT1alpha, MEF2, c-Myc, ATF-2 and c-Fos were induced early during infection, and ERK inhibition significantly blocked the c-Fos, c-Jun, c-Myc, and STAT1alpha activation in the infected cells. AP1 transcription factors binding to the RTA promoter in electrophoretic mobility shift assays were readily detected in the infected cell nuclear extracts which were significantly reduced by ERK inhibition. Together, these results suggest that very early during de novo infection, KSHV induces the ERK1/2 to modulate the initiation of viral gene expression and host cell genes, which further supports our hypothesis that beside the conduit for viral DNA delivery into the cytoplasm, KSHV interactions with host cell receptor(s) create an appropriate intracellular environment facilitating infection.
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PMID:ERK1/2 and MEK1/2 induced by Kaposi's sarcoma-associated herpesvirus (human herpesvirus 8) early during infection of target cells are essential for expression of viral genes and for establishment of infection. 1605 24

Despite their benign histological appearance, juvenile angiofibromas sometimes exhibit an aggressive growth behavior. Molecular and genetic analyses have detected beta-catenin mutations and androgen receptor gene gains in this tumor. Because intensive cross-talk among beta-catenin, androgen receptor, and C-MYC has been detected recently, we analyzed expression of the C-MYC protooncogene (MYC) on the genetic, transcriptional and translational level in seven sporadic juvenile angiofibromas. Two-color in situ hybridization analyses for chromosome 8 and MYC found in all seven juvenile angiofibromas significant MYC losses. In the three advanced juvenile angiofibromas of this series (Fisch stages III and IV) additional significant MYC gains were observed demonstrating a genetic heterogeneity for the MYC protooncogene. In cases of genetic MYC heterogeneity, reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, Western blot investigations, and immunohistology showed increased C-MYC mRNA and protein levels. Semiquantitative RT-PCR analyses from laser microdissected endothelial cells and fibroblasts found no differences of C-MYC mRNA levels, leaving open the question of the neoplastic cell in juvenile angiofibromas. The finding of genetic MYC heterogeneity associated with C-MYC overexpression on the mRNA and protein level in advanced juvenile angiofibromas indicates involvement of the MYC oncogene in aggressive growth behavior.
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PMID:Genetic heterogeneity of the MYC oncogene in advanced juvenile angiofibromas. 1636 59

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

Chromosomal aberrations (amplifications and deletions) underlie the genesis or development of cancer. Amplification of 8q24 is one of the most frequent events in esophageal cancer. To define whether C-MYC is the target gene for 8q24 amplification, we performed fluorescence in situ hybridization using a MYC (8q24.12 approximately q24.13) probe in esophageal cancer from southern China. Furthermore, we detected the expression status of several genes including C-MYC, TRIB1 (alias C8FW), and FAM84B (alias NSE2) in the regions of 8q24 via reverse transcriptase-polymerase chain reaction or immunohistochemical analysis (or both). Distinct amplification of 8q24 was found in esophageal carcinomas. Only 4 of 46 cases showed obvious protein expression in part of the esophageal cancerous nest. In particular, increased protein expression of C-MYC was shown only in a small part of a cancerous nest in the four cases. Positive C-MYC staining was detected mainly in the cytoplasm of esophageal cancer cells. No expression of TRIB1 was detected in esophageal squamous cell carcinomas. Of 59 cases, 39 (66%) cases showed increased expression of FAM84B in esophageal carcinomas. The results suggest that C-MYC and TRIB1 may not be the amplification target of 8q24 in esophageal cancer. FAM84B might be involved in the genesis or development of esophageal cancer in southern China. Whether FAM84B is the amplification target of esophageal cancer awaits further investigation.
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PMID:Negative implication of C-MYC as an amplification target in esophageal cancer. 1649 May 93


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