Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing, China.
Nucleic Acids Res. 2020 Apr 17;48(7):3619-3637. doi: 10.1093/nar/gkaa096.
REV3L, the catalytic subunit of DNA polymerase ζ (Pol ζ), is indispensable for translesion DNA synthesis, which protects cells from deleterious DNA lesions resulting from various intrinsic and environmental sources. However, REV3L lacks a proofreading exonuclease activity and consequently bypasses DNA lesions at the expense of increased mutations, which poses a severe threat to genome stability. Here we report a site-specific proteolytic event of human REV3L. We show that REV3L is cleaved by a threonine aspartase, Taspase1 (TASP1), to generate an N-terminal 70-kDa fragment (N70) and a polypeptide carrying the C-terminal polymerase catalytic domain in human cells. Strikingly, such a post-translational cleavage event plays a vital role in controlling REV3L stability by preventing ubiquitination and proteasome-mediated degradation of REV3L. Indicative of the biological importance of the above REV3L post-translational processing, cellular responses to UV and cisplatin-induced DNA lesions are markedly impaired in human HCT116 cell derivatives bearing defined point mutations in the endogenous REV3L gene that compromise REV3L cleavage. These findings establish a new paradigm in modulating the abundance of REV3L through site-specific proteolysis in human cells.
REV3L 是 DNA 聚合酶 ζ (Pol ζ) 的催化亚基，对跨损伤 DNA 合成至关重要，这种合成可以保护细胞免受各种内在和环境来源的有害 DNA 损伤。然而，REV3L 缺乏校对外切酶活性，因此会在牺牲增加突变的情况下绕过 DNA 损伤，这对基因组稳定性构成了严重威胁。在这里，我们报告了人类 REV3L 的一个位点特异性蛋白水解事件。我们表明，REV3L 被苏氨酸天冬氨酸酶 Taspase1 (TASP1) 切割，生成一个 70 kDa 的 N 端片段 (N70) 和一个带有 C 端聚合酶催化结构域的多肽在人类细胞中。引人注目的是，这种翻译后切割事件通过防止 REV3L 的泛素化和蛋白酶体介导的降解，对控制 REV3L 稳定性起着至关重要的作用。上述 REV3L 翻译后加工的生物学重要性的指示，是在携带内源性 REV3L 基因中定义点突变的人 HCT116 细胞衍生物中，细胞对 UV 和顺铂诱导的 DNA 损伤的反应明显受损，这些突变会损害 REV3L 的切割。这些发现为通过人类细胞中的位点特异性蛋白水解来调节 REV3L 的丰度建立了一个新的范例。
