Department of Surgery, Johns Hopkins University, Baltimore, Maryland.
Department of Molecular Biology and Genetics, Johns Hopkins University, Baltimore, Maryland.
Genes Chromosomes Cancer. 2019 Aug;58(8):530-540. doi: 10.1002/gcc.22735. Epub 2019 Feb 10.
Telomerase reverse transcriptase (TERT) activation plays an important role in cancer development by enabling the immortalization of cells. TERT regulation is multifaceted, and its promoter methylation has been implicated in controlling expression through alteration in transcription factor binding. We have characterized TERT promoter methylation, transcription factor binding, and TERT expression levels in five differentiated thyroid cancer (DTC) cell lines and six normal thyroid tissue samples by targeted bisulfite sequencing, ChIP-qPCR, and qRT-PCR. DTC cell lines express varying levels of TERT and exhibit TERT promoter methylation patterns similar to patterns seen in other telomerase positive cancer cell lines. The minimal promoter immediately surrounding the transcription start site is hypomethylated, while further upstream portions show dense methylation. In contrast, the TERT promoter in normal thyroid tissue is largely unmethylated throughout and expresses TERT minimally. Transcription factor binding is also affected by TERT mutation status. The E-twenty-six (ETS) factor GABPA exhibits TERT binding in the TERT mutant DTC cells only, and allele-specific methylation patterns at the minimal promoter were observed as well, which may indicate allele-specific factor recruitment at the minimal promoter. Furthermore, we identified binding sites for activators MYC and GSC in the hypermethylated upstream region, pointing to its possible importance in TERT regulation. Overall, TERT expression and telomerase activity depend on the interplay of multiple regulatory mechanisms including TERT promoter methylation, mutation status, and recruitment of transcription factors. This work explores of the interplay between these regulatory mechanisms and offers insight into cellular control of active telomerase in human cancer.
端粒酶逆转录酶（TERT）的激活在癌症发展中起着重要作用，使细胞实现永生化。TERT 的调节具有多面性，其启动子甲基化通过改变转录因子结合来控制表达。我们通过靶向亚硫酸氢盐测序、ChIP-qPCR 和 qRT-PCR 对 5 种分化型甲状腺癌（DTC）细胞系和 6 份正常甲状腺组织样本中的 TERT 启动子甲基化、转录因子结合和 TERT 表达水平进行了表征。DTC 细胞系表达不同水平的 TERT，并表现出与其他端粒酶阳性癌细胞系相似的 TERT 启动子甲基化模式。转录起始位点周围的最小启动子呈低甲基化，而上游部分则呈密集甲基化。相比之下，正常甲状腺组织中的 TERT 启动子在整个区域都呈低甲基化状态，表达的 TERT 较少。转录因子结合也受 TERT 突变状态的影响。ETS 因子 GABPA 在 TERT 突变的 DTC 细胞中仅表现出 TERT 结合，并且在最小启动子上观察到等位基因特异性甲基化模式，这可能表明最小启动子上的等位基因特异性因子募集。此外，我们在高度甲基化的上游区域中鉴定到了激活子 MYC 和 GSC 的结合位点，表明其在 TERT 调节中的可能重要性。总的来说，TERT 表达和端粒酶活性取决于多个调节机制的相互作用，包括 TERT 启动子甲基化、突变状态和转录因子的募集。这项工作探讨了这些调节机制之间的相互作用，并深入了解了人类癌症中活跃端粒酶的细胞控制。
