Molecular studies on mammalian telomerase

端粒酶是一种核糖核蛋白复合体，可以重新合成富含端粒g的DNA链。它是一种特殊的逆转录酶，包含催化组分TERT和模板RNA tr。为了更好地了解端粒酶的调控机制，在本研究中，我们首先建立了TERT^<-1->敲除小鼠。第一代TERT^<-1->基因敲除小鼠未表现出任何表型，表明端粒酶催化蛋白本身缺失不影响小鼠发育。然而，当TERT^<-1->小鼠进行交配以保持端粒酶缺失时，第五代小鼠的生殖系细胞明显萎缩。这些结果表明生殖细胞对端粒长度的减少特别敏感。我们和其他人已经证明，TERT基因表达水平在调节细胞端粒酶活性方面起主要作用。尽管有报道称，包括Myc、Sp1、ER在内的几种转录因子参与了TERT的表达，但关于TERT的具体调控机制，特别是在正常体细胞中，仍然知之甚少。我们研究了正常的人外周血淋巴细胞作为模型系统。我们发现转录因子USF1和2在体外正调控TERT启动子。截断形式的USF1和2，迷你USF1和迷你usf2，已知是由相同的基因通过选择性剪接产生的。有趣的是，我们在体外证明了mini-USF1和2负调控启动子。人外周血淋巴细胞大多处于静止状态，不具有端粒酶活性。当外部刺激作用于静止的淋巴细胞时，它们开始增殖，同时开始显示端粒酶活性。我们发现负调节因子mini-USFs存在于静息细胞中，而不存在于活化淋巴细胞中。相反，在静止和生长的淋巴细胞中均存在正调节因子全长usf。这些结果表明，在控制TERT表达方面，迷你usf可能对完整usf有主要的负作用。少

Telomerase is a ribonucleoprotein complex that synthesizes telomeric G-rich strand DNA de novo. It is a specialized reverse transcriptase, containing the catalytic component, TERT and the template RNA, TR.To better understand regulation mechanisms of telomerase, in this study, we first established TERT^<-1-> knockout mice. The first generation of TERT^<-1-> knockout mice did not show any phenotype, indicating that the absence of telomerase catalytic protein by itself does not affect mouse development. However, when TERT^<-1-> mice were mated to keep the line absent for telomerase, the fifth generation mice showed significantly hypotrophic germ line cells. These results indicate that germ cells are particularly sensitive to telomere length reduction. We and others have already demonstrated that TERT gene expression levels have a primary role in regulating telomerase activity in cells. Although it has been reported that several transcription factors, including Myc, Sp1, ER, are involved … More in TERT expression, details of mechanisms regulating TERT, especially in normal somatic cells, remain largely unknown. We have studied normal human peripheral lymphocytes as a model system. We found that the transcription factors USF1 and 2 positively regulate the TERT promoter in vitro. Truncated forms of USF1 and 2, mini-USF1 and mini-USF2, are known to be produced from the same genes by alternative splicing. Interestingly, we demonstrated that mini-USF1 and 2 negatively regulate the promoter in vitro. Human peripheral lymphocytes are mostly resting and do not possess telomerase activity. Upon external stimuli applied to resting lymphocytes, they start proliferation and simultaneously begin to show telomerase activity. We found that the negative regulator, mini-USFs are present in resting cells but not in activated lymphocytes. In contrast, the positive regulator, full length USFs are present both in resting and growing lymphocytes. These results suggest that mini-USFs might have dominant negative effects on full USFs in controlling TERT expression. Less