scholarly journals Latency-Associated Nuclear Antigen of Kaposi's Sarcoma-Associated Herpesvirus Up-Regulates Transcription of Human Telomerase Reverse Transcriptase Promoter through Interaction with Transcription Factor Sp1

2004 ◽  
Vol 78 (19) ◽  
pp. 10348-10359 ◽  
Author(s):  
Subhash C. Verma ◽  
Sumit Borah ◽  
Erle S. Robertson
Keyword(s):  
Reverse Transcriptase ◽  
Kaposi's Sarcoma ◽  
Promoter Activity ◽  
Human Cells ◽  
Nuclear Antigen ◽  
Telomerase Reverse Transcriptase ◽  
Htert Promoter ◽  
Human Telomerase Reverse Transcriptase ◽  
Infected Cells ◽  
Human Telomerase

ABSTRACT Telomerase is required for the maintenance of telomere length and is an important determinant for cell immortalization. In human cells, telomerase activity is due to the expression of its enzymatic subunit, human telomerase reverse transcriptase (hTERT). The expression of hTERT is not typically detectable in healthy somatic human cells but is present in cancerous tissues and immortalized cells. We have previously shown that hTERT promoter activity is up-regulated by the Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded latency-associated nuclear antigen (LANA). LANA is expressed in all forms of human malignancies associated with KSHV. The hTERT promoter sequence located at positions −130 to +5 contains several Sp1 binding motifs and was shown be important for up-regulation by LANA. In this report, we demonstrate that hTERT promoter activity is due to the direct interaction of LANA with Sp1. The interaction of LANA with Sp1 was demonstrated through in vitro binding experiments and coimmunoprecipitation and is supported by the colocalization of these two molecules in the nuclei of KSHV-infected cells. Moreover, LANA modulates Sp1-mediated transcription in transient GAL4 fusion reporter assays. Mapping of the regions involved in binding and transcriptional activation showed that the amino terminus of LANA is the major site for interaction and up-regulation but that it can cooperate with the carboxy terminus to enhance these functions. An analysis of Sp1 binding to its cognate sequence corroborated the binding data. Together, our results suggest that the interaction of LANA with Sp1 up-regulates the telomerase promoter, potentially contributing to the immortalization of KSHV-infected cells.

Effect of RNA interference targeting human telomerase reverse transcriptase on telomerase and its related protein expression in nasopharyngeal carcinoma cells

2006 ◽  
Vol 121 (5) ◽  
pp. 476-482 ◽  
Author(s):  
Yan Wang ◽  
Hong-Gang Duan ◽  
Shi-Ming Chen ◽  
Bo-Kui Xiao ◽  
Jie Cheng ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Proliferating Cell Nuclear Antigen ◽  
Short Hairpin Rna ◽  
Nuclear Antigen ◽  
Telomerase Reverse Transcriptase ◽  
Hairpin Rna ◽  
Human Telomerase Reverse Transcriptase ◽  
Short Hairpin ◽  
Human Telomerase ◽  
Proliferating Cell

Objective: Analysis of the correlation between telomerase and the expression of its related proteins may provide insight into the molecular mechanism of nasopharyngeal carcinogenesis. We investigated the effect of short hair pin ribonucleic acid (RNA) specific for human telomerase reverse transcriptase messenger RNA on the expression of the proteins c-myc (the transcription factor c-myc is a shortlived nuclear phospho-protein involved in cell proliferation and differentiation, belongs to the myc family), proliferating cell nuclear antigen and Caspase-3 in nasopharyngeal carcinoma cells.Methods: Short hairpin RNA expression vectors targeting the messenger RNA of human telomerase reverse transcriptase were constructed. Cells were treated with the short hairpin RNA expression vectors targeting human telomerase reverse transcriptase or vectors that included mismatched short hairpin RNA, and telomerase activity was measured by telomeric repeat amplification enzyme-linked immunosorbent assay. Cell viability was examined using the 3-(4,5-dimethyl thizol-2-yl) 2,5-diphenyl tetrazolium bromide assay. The expression of the three proteins (c-myc, proliferating cell nuclear antigen and Caspase-3) was determined by Western blotting.Results: Short hairpin RNA specific for human telomerase reverse transcriptase messenger RNA significantly inhibited telomerase activity. In addition, the expression of and proliferating cell nuclear antigen were both inhibited, while the expression of Caspase-3 was up-regulated.Conclusions: Our results suggest that short hairpin RNA directed against human telomerase reverse transcriptase inhibits cell viability by regulating telomerase activity and its related proteins expression in nasopharyngeal carcinoma cells. Therefore, RNA interference technology may be a promising strategy for the treatment of nasopharyngeal cancer.

scholarly journals Prognostic significance of human telomerase reverse transcriptase promoter region mutations C228T and C250T for overall survival in spinal chordomas

2019 ◽  
Vol 21 (8) ◽  
pp. 1005-1015 ◽  
Author(s):  
Chetan Bettegowda ◽  
Stephen Yip ◽  
Bowen Jiang ◽  
Wei-Lien Wang ◽  
Michelle J Clarke ◽  
...  
Keyword(s):  
Overall Survival ◽  
Local Recurrence ◽  
Reverse Transcriptase ◽  
Prognostic Significance ◽  
Spinal Tumors ◽  
Telomerase Reverse Transcriptase ◽  
Htert Promoter ◽  
Human Telomerase Reverse Transcriptase ◽  
Mutational Status ◽  
Human Telomerase

Abstract Background Spinal chordomas, a subtype of primary spinal column malignancies (PSCM), are rare tumors with poor prognosis, and we have limited understanding of the molecular drivers of neoplasia. Methods Study design was a retrospective review of prospectively collected data with cross-sectional survival. Archived paraffin embedded pathologic specimens were collected for 133 patients from 6 centers within Europe and North America between 1987 and 2012. Tumor DNA was extracted and the human telomerase reverse transcriptase (hTERT) promoter was sequenced. The hTERT mutational status was correlated with overall survival (OS) and time to first local recurrence. Results Ninety-two chordomas, 26 chondrosarcomas, 7 osteosarcomas, 3 Ewing’s sarcomas, and 5 other malignant spinal tumors were analyzed. Median OS following surgery was 5.8 years (95% CI: 4.6 to 6.9) and median time to first local recurrence was 3.9 years (95% CI: 2.5 to 6.7). Eight chordomas, 2 chondrosarcomas, 1 Ewing’s sarcoma, and 1 other malignant spinal tumor harbored either a C228T or C250T mutation in the hTERT promoter. In the overall cohort, all patients with hTERT mutation were alive at 10 years postoperative with a median OS of 5.1 years (95% CI: 4.5 to 6.6) (P = 0.03). hTERT promoter mutation was observed in 8.7% of spinal chordomas, and 100% of chordoma patients harboring the mutation were alive at 10 years postoperative compared with 67% patients without the mutation (P = 0.05). Conclusions We report for the first time that hTERT promoter mutations C228T and C250T are present in approximately 8.7% of spinal chordomas. The presence of hTERT mutations conferred a survival benefit and could potentially be a valuable positive prognostic molecular marker in spinal chordomas.

scholarly journals Telomerase Activation by Human Papillomavirus Type 16 E6 Protein: Induction of Human Telomerase Reverse Transcriptase Expression through Myc and GC-Rich Sp1 Binding Sites

2001 ◽  
Vol 75 (12) ◽  
pp. 5559-5566 ◽  
Author(s):  
Stephen T. Oh ◽  
Saturo Kyo ◽  
Laimonis A. Laimins
Keyword(s):  
Reverse Transcriptase ◽  
Binding Sites ◽  
Telomerase Activity ◽  
Telomerase Reverse Transcriptase ◽  
Htert Promoter ◽  
Htert Expression ◽  
Human Telomerase Reverse Transcriptase ◽  
Htert Transcription ◽  
Human Telomerase ◽  
In Cells

ABSTRACT High-risk human papillomaviruses (HPVs) immortalize keratinocytes by disrupting the retinoblastoma protein (Rb)/p16 pathway and activating telomerase. The E7 oncoprotein targets Rb, while the E6 oncoprotein induces telomerase activity in human keratinocytes. This study has examined the mechanism by which E6 activates telomerase. Expression of human telomerase reverse transcriptase (hTERT), the catalytic subunit of telomerase, was found to be increased in keratinocytes stably expressing HPV type 16 E6, suggesting that E6 acts to increase hTERT transcription. hTERT expression and telomerase activity were activated to significantly higher levels in cells expressing both E6 and E7 than in cells expressing E6 alone. This indicates that E7 may augment E6-mediated activation of hTERT transcription. In transient-transfection assays using hTERT reporters, the induction of hTERT expression by E6 was found to be mediated by a 258-bp fragment of the hTERT promoter, proximal to the ATG initiation codon. Previous studies have demonstrated that overexpression of Myc can activate hTERT expression, suggesting that Myc may be a mediator of E6-mediated hTERT induction. However, in cells stably expressing E6, no strict correlation between the level of Myc and the activation of hTERT was found. Consistent with this observation, mutation of the two Myc binding sites in the hTERT promoter only modestly reduced responsiveness to E6 in transient reporter assays. This indicates that activation of Myc-dependent transcription is not essential for E6-mediated upregulation of hTERT expression. The hTERT promoter also contains five GC-rich elements that can bind Sp1. Mutation of these sites within the 258-bp fragment partially reduced hTERT induction by E6. However, when mutations in the Sp1 sites were combined with the mutated Myc binding sites, all activation by E6 was lost. This indicates that it is the combinatorial binding of factors to Myc and Sp1 cis elements that is responsible for hTERT induction by E6.

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