Sp1 and Sp3 Recruit Histone Deacetylase to Repress Transcription of Human Telomerase Reverse Transcriptase (hTERT) Promoter in Normal Human Somatic Cells

Jaejoon Won,Jeongbin Yim,Tae Kook Kim

doi:10.1074/jbc.m206064200

Abstract

Activation of telomerase is crucial for cells to gain immortality. In human cells, telomerase activity is tightly regulated by the expression of its catalytic subunit, human telomerase reverse transcriptase (hTERT). In most normal human somatic cells, hTERT is not expressed, and its suppression acts as an important gatekeeper against tumorigenesis. Here we describe the systematic analyses of hTERT promoter to understand the transcriptional repression mechanism of the hTERT gene in normal human somatic cells. Through the serial deletion analysis of hTERT promoter in normal human fibroblasts, we identified a critical repressive element on the hTERT promoter. The repressive element formed DNA-protein complexes with Sp1 and Sp3 in nuclear extracts. Using formaldehyde cross-linked chromatin immunoprecipitation analysis, we found that Sp1 and Sp3 were associated with the endogenously repressed hTERT promoter in human fibroblasts. Furthermore, Sp1 and Sp3 interacted with histone deacetylase (HDAC) in these cells. Overexpression of dominant-negative mutants of Sp1 and Sp3, which contained mainly the HDAC2-binding domain, relieved the HDAC-mediated repression of the hTERT promoter. Taken together, these results suggest that Sp1 and Sp3 associate with the hTERT promoter, recruiting HDAC for the localized deacetylation of nucleosomal histones and transcriptional silencing of the hTERT gene in normal human somatic cells.

Highlights

Activation of telomerase is crucial for cells to gain immortality
Overexpression of dominant-negative mutants of Sp1 and Sp3, which contained mainly the HDAC2-binding domain, relieved the histone deacetylase (HDAC)-mediated repression of the human telomerase reverse transcriptase (hTERT) promoter. These results suggest that Sp1 and Sp3 associate with the hTERT promoter, recruiting HDAC for the localized deacetylation of nucleosomal histones and transcriptional silencing of the hTERT gene in normal human somatic cells
P-188 had less activity than p-179 in the absence of trichostatin A (TSA) treatment. These results suggest that the region between Ϫ188 and Ϫ180 of the hTERT promoter may be engaged in the HDAC-mediated tight transcriptional repression of the hTERT gene in normal human somatic cells

Summary

Introduction

Activation of telomerase is crucial for cells to gain immortality. In human cells, telomerase activity is tightly regulated by the expression of its catalytic subunit, human telomerase reverse transcriptase (hTERT). Further deletion from Ϫ274 to Ϫ189 caused a slight increase in promoter activity, consistent with our previous report that the E-box within this region acts as the repressive element through interaction with the Mad-Max complex in normal human somatic cells [19].

Results

Conclusion