Human Telomerase Reverse Transcriptase (hTERT) Transcription Requires Sp1/Sp3 Binding to the Promoter and a Permissive Chromatin Environment
Human Telomerase Reverse Transcriptase (hTERT) Transcription Requires Sp1/Sp3 Binding to the Promoter and a Permissive Chromatin Environment
复制标题
DOI：
10.1074/jbc.m115.662221
复制
发表时间：
2015-12-11
影响因子：
4.8
通讯作者：
Zhu, Jiyue
中科院分区：
文献类型：
--
作者：
Cheng, De;Zhao, Yuanjun;Zhu, Jiyue
人类端粒酶基因hTERT的转录受转录因子（包括Sp1家族蛋白）及其染色质环境的调控。为了了解其在相关染色质背景下的调控，我们使用了含有160 kb含有hTERT基因的人类基因组序列的细菌人工染色体报告器。在染色体整合后，细菌人工染色体重述内源性hTERT表达，与瞬时报告者相反。Sp1/Sp3的表达与hTERT启动子活性无关，在端粒酶阳性和端粒酶阴性细胞中，这些tf与hTERT启动子结合。近端GC-box突变导致hTERT启动子活性显著降低，5个GC-box突变均消除了其转录活性。在端粒酶阳性和阴性细胞中，gc -box的突变和内源性Sp1的敲低都不会影响启动子与其他tf的结合，包括e- box结合蛋白，以及hTERT启动子上组蛋白H3K9的乙酰化和三甲基化。结果表明，Sp1/Sp3启动子结合是hTERT转录的必要步骤，但不是限制性步骤。hTERT转录需要一个允许的染色质环境。重要的是，我们的数据还揭示了gc -box和e -box在htert调控中的不同功能；虽然gc -box对启动子活性至关重要，但与e- box结合的因子可以抑制hTERT启动子。
The transcription of human telomerase gene hTERT is regulated by transcription factors (TFs), including Sp1 family proteins, and its chromatin environment. To understand its regulation in a relevant chromatin context, we employed bacterial artificial chromosome reporters containing 160 kb of human genomic sequence containing the hTERT gene. Upon chromosomal integration, the bacterial artificial chromosomes recapitulated endogenous hTERT expression, contrary to transient reporters. Sp1/Sp3 expression did not correlate with hTERT promoter activity, and these TFs bound to the hTERT promoters in both telomerase-positive and telomerase-negative cells. Mutation of the proximal GC-box resulted in a dramatic decrease of hTERT promoter activity, and mutations of all five GC-boxes eliminated its transcriptional activity. Neither mutations of GC-boxes nor knockdown of endogenous Sp1 impacted promoter binding by other TFs, including E-box-binding proteins, and histone acetylation and trimethylation of histone H3K9 at the hTERT promoter in telomerase-positive and -negative cells. The result indicated that promoter binding by Sp1/Sp3 was essential, but not a limiting step, for hTERT transcription. hTERT transcription required a permissive chromatin environment. Importantly, our data also revealed different functions of GC-boxes and E-boxes in hTERT-regulation; although GC-boxes were essential for promoter activity, factors bound to the E-boxes functioned to de-repress hTERT promoter.