Chiba Kunitoshi, Vogan Jacob M, Wu Robert A, Gill Manraj S, Zhang Xiaozhu, Collins Kathleen, Hockemeyer Dirk
Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA.
Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA
Mol Cell Biol. 2017 Jan 19;37(3). doi: 10.1128/MCB.00541-16. Print 2017 Feb 1.
Telomerase action at telomeres is essential for the immortal phenotype of stem cells and the aberrant proliferative potential of cancer cells. Insufficient telomere maintenance can cause stem cell and tissue failure syndromes, while increased telomerase levels are associated with tumorigenesis. Both pathologies can arise from only small perturbation of telomerase function. To analyze telomerase at its low endogenous expression level, we genetically engineered human pluripotent stem cells (hPSCs) to express various N-terminal fusion proteins of the telomerase reverse transcriptase from its endogenous locus. Using this approach, we found that these modifications can perturb telomerase function in hPSCs and cancer cells, resulting in telomere length defects. Biochemical analysis suggests that this defect is multileveled, including changes in expression and activity. These findings highlight the unknown complexity of telomerase structural requirements for expression and function in vivo.
端粒酶在端粒处的作用对于干细胞的永生化表型和癌细胞异常的增殖潜能至关重要。端粒维持不足可导致干细胞和组织衰竭综合征，而端粒酶水平升高与肿瘤发生相关。这两种病理情况都可能仅由端粒酶功能的微小扰动引起。为了在其低内源性表达水平下分析端粒酶，我们对人类多能干细胞（hPSC）进行基因工程改造，使其从内源基因座表达端粒酶逆转录酶的各种N端融合蛋白。使用这种方法，我们发现这些修饰可扰乱hPSC和癌细胞中的端粒酶功能，导致端粒长度缺陷。生化分析表明，这种缺陷是多层面的，包括表达和活性的变化。这些发现突出了端粒酶在体内表达和功能的结构要求方面未知的复杂性。
