Characterization of the telomerase modulating activities of yeast DNA helicases.
Characterization of the telomerase modulating activities of yeast DNA helicases.
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DOI：
10.1016/bs.mie.2021.08.005
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发表时间：
2021
影响因子：
--
通讯作者：
Bochman ML
中科院分区：
文献类型：
--
作者：
Nickens DG;Bochman ML
具有线性染色体的真核生物通过一种称为端粒酶的专门的核糖核蛋白逆转录酶的作用来规避末端复制问题。缺乏端粒酶活性的细胞在染色体末端缩短到临界长度时会衰老。相反，癌细胞可以通过在DNA复制的每个周期中上调端粒酶以延长端粒而永生化。因此，端粒酶的调节对于正常的端粒稳态是至关重要的。在体内调节端粒酶活性的各种已知方式中，最近的研究表明DNA解旋酶参与其中。在酿酒酵母中，Hrq1和Pif1解旋酶在调节端粒和DNA双链DNA断裂处的端粒酶延伸的途径中起作用。体外分析表明，当这些解旋酶在反应中组合时，它们协同抑制或刺激端粒酶活性，这取决于哪种解旋酶具有催化活性。在这里，我们描述了Hrq 1和Pif 1的过量生产和纯化方法。我们还报告了制备部分纯化的细胞提取物与端粒酶活性和如何在体外这些解旋酶对端粒酶活性的影响可以进行评估。
Eukaryotes with linear chromosomes circumvent the end replication problem via the action of a specialized ribonucleoprotein reverse transcriptase known as telomerase. Cells lacking telomerase activity will senesce when their chromosome ends shorten to a critical length. In contrast, cancer cells can become immortalized by upregulating telomerase to lengthen telomeres during each cycle of DNA replication. Thus, the regulation of telomerase is critical for normal telomere homeostasis. Of the various known ways that telomerase activity is modulated in vivo, recent studies have demonstrated that DNA helicases are involved. In Saccharomyces cerevisiae, the Hrq1 and Pif1 helicases act in a pathway that regulates telomerase extension at telomeres and at DNA double-strand DNA breaks. In vitro analysis demonstrates that when these helicases are combined in reactions, they synergistically inhibit or stimulate telomerase activity depending on which helicase is catalytically active. Here, we describe the methods for the overproduction and purification of Hrq1 and Pif1. We also report the preparation of partially purified cell extracts with telomerase activity and how the effects of these helicase on telomerase activity can be assessed in vitro.
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DOI：
10.1038/nrg3296
发表时间：
2012-11
期刊：
Nature reviews. Genetics
影响因子：
--
作者：
通讯作者：
--
影响因子：
3.8
作者：
Bochman ML;Sabouri N;Zakian VA
通讯作者：
Zakian VA
影响因子：
9.2
作者：
Huang, PH;Pryde, FE;Louis, EJ
通讯作者：
Louis, EJ
影响因子：
2.9
作者：
Andis, Nicholas M.;Sausen, Christopher W.;Bochman, Matthew L.
通讯作者：
Bochman, Matthew L.
影响因子：
64.8
作者：
Boulé, JB;Vega, LR;Zakian, VA
通讯作者：
Zakian, VA