Department of Human Genetics, Leiden University Medical Center, Einthovenweg 20, 2333 ZC, Leiden, The Netherlands.
Sci Rep. 2020 Mar 9;10(1):4332. doi: 10.1038/s41598-020-61243-4.
Transcription-coupled repair (TCR) removes DNA lesions from the transcribed strand of active genes. Stalling of RNA polymerase II (RNAPII) at DNA lesions initiates TCR through the recruitment of the CSB and CSA proteins. The full repertoire of proteins required for human TCR - particularly in a chromatin context - remains to be determined. Studies in mice have revealed that the nucleosome-binding protein HMGN1 is required to enhance the repair of UV-induced lesions in transcribed genes. However, whether HMGN1 is required for human TCR remains unaddressed. Here, we show that knockout or knockdown of HMGN1, either alone or in combination with HMGN2, does not render human cells sensitive to UV light or Illudin S-induced transcription-blocking DNA lesions. Moreover, transcription restart after UV irradiation was not impaired in HMGN-deficient cells. In contrast, TCR-deficient cells were highly sensitive to DNA damage and failed to restart transcription. Furthermore, GFP-tagged HMGN1 was not recruited to sites of UV-induced DNA damage under conditions where GFP-CSB readily accumulated. In line with this, HMGN1 did not associate with the TCR complex, nor did TCR proteins require HMGN1 to associate with DNA damage-stalled RNAPII. Together, our findings suggest that HMGN1 and HMGN2 are not required for human TCR.
转录偶联修复（TCR）可从活跃基因的转录链上去除 DNA 损伤。RNA 聚合酶 II（RNAPII）在 DNA 损伤处的停滞通过 CSB 和 CSA 蛋白的募集来启动 TCR。人类 TCR 所需的全套蛋白质 - 尤其是在染色质背景下 - 仍有待确定。在小鼠中的研究表明，核小体结合蛋白 HMGN1 对于增强转录基因中 UV 诱导损伤的修复是必需的。然而，HMGN1 是否是人类 TCR 的必需蛋白尚未得到解决。在这里，我们表明，单独或与 HMGN2 联合敲除或敲低 HMGN1 不会使人类细胞对 UV 光或 Illudin S 诱导的转录阻断 DNA 损伤敏感。此外，UV 照射后转录的重新启动在 HMGN 缺陷细胞中并未受损。相比之下，TCR 缺陷细胞对 DNA 损伤高度敏感，并且无法重新启动转录。此外，在 GFP-CSB 容易积累的条件下，GFP 标记的 HMGN1 未被募集到 UV 诱导的 DNA 损伤部位。与此一致的是，HMGN1 不与 TCR 复合物相关联，并且 TCR 蛋白也不需要 HMGN1 与 DNA 损伤暂停的 RNAPII 相关联。总之，我们的研究结果表明，HMGN1 和 HMGN2 对于人类 TCR 不是必需的。
