Department of Biological Sciences, University of Chicago, Chicago, IL 60637, USA.
SynRx Therapeutics, Chicago, IL 60642, USA.
Front Biosci (Landmark Ed). 2024 Jul 23;29(7):262. doi: 10.31083/j.fbl2907262.
The switching/sucrose non-fermentable (SWI/SNF) Related, Matrix Associated, Actin Dependent Regulator Of Chromatin, Subfamily A (SMARCA) member 2 and member 4 (SMARCA2/4) are paralogs and act as the key enzymatic subunits in the SWI/SNF complex for chromatin remodeling. However, the role of SMARCA2/4 in DNA damage response remains unclear.
Laser microirradiation assays were performed to examine the key domains of SMARCA2/4 for the relocation of the SWI/SNF complex to DNA lesions. To examine the key factors that mediate the recruitment of SMARCA2/4, the relocation of SMARCA2/4 to DNA lesions was examined in HeLa cells treated with inhibitors of Ataxia-telangiectasia-mutated (ATM), Ataxia telangiectasia and Rad3-related protein (ATR), CREB-binding protein (CBP) and its homologue p300 (p300/CBP), or Poly (ADP-ribose) polymerase (PARP) 1/2 as well as in H2AX-deficient HeLa cells. Moreover, by concomitantly suppressing SMARCA2/4 with the small molecule inhibitor FHD286 or Compound 14, the function of SMARCA2/4 in Radiation sensitive 51 (RAD51) foci formation and homologous recombination repair was examined. Finally, using a colony formation assay, the synergistic effect of PARP inhibitors and SMARCA2/4 inhibitors on the suppression of tumor cell growth was examined.
We show that SMARCA2/4 relocate to DNA lesions in response to DNA damage, which requires their ATPase activities. Moreover, these ATPase activities are also required for the relocation of other subunits in the SWI/SNF complex to DNA lesions. Interestingly, the relocation of SMARCA2/4 is independent of γH2AX, ATM, ATR, p300/CBP, or PARP1/2, indicating that it may directly recognize DNA lesions as a DNA damage sensor. Lacking SMARCA2/4 prolongs the retention of γH2AX, Ring Finger Protein 8 (RNF8) and Breast cancer susceptibility gene 1 (BRCA1) at DNA lesions and impairs RAD51-dependent homologous recombination repair. Furthermore, the treatment of an SMARCA2/4 inhibitor sensitizes tumor cells to PARP inhibitor treatment.
This study reveals SMARCA2/4 as a DNA damage repair factor for double-strand break repair.
开关/蔗糖非发酵（SWI/SNF）相关、基质相关、肌动蛋白依赖性染色质调节剂亚家族 A（SMARCA）成员 2 和成员 4（SMARCA2/4）是同源物，作为染色质重塑 SWI/SNF 复合物的关键酶亚基。然而，SMARCA2/4 在 DNA 损伤反应中的作用尚不清楚。
激光微照射实验用于研究 SMARCA2/4 的关键结构域，以研究 SWI/SNF 复合物向 DNA 损伤的重定位。为了研究介导 SMARCA2/4 募集的关键因素，在使用 ATM 抑制剂、ATR 抑制剂、CBP 及其同源物 p300（p300/CBP）、或聚（ADP-核糖）聚合酶 1/2（PARP1/2）抑制剂处理的 HeLa 细胞中以及在 H2AX 缺陷型 HeLa 细胞中，研究了 SMARCA2/4 向 DNA 损伤的重定位。此外，通过同时用小分子抑制剂 FHD286 或化合物 14 抑制 SMARCA2/4，研究了 SMARCA2/4 在 RAD51 焦点形成和同源重组修复中的功能。最后，通过集落形成实验，研究了 PARP 抑制剂和 SMARCA2/4 抑制剂对肿瘤细胞生长抑制的协同作用。
我们表明，SMARCA2/4 在 DNA 损伤后会重新定位到 DNA 损伤部位，这需要它们的 ATP 酶活性。此外，这些 ATP 酶活性对于 SWI/SNF 复合物的其他亚基向 DNA 损伤的重定位也是必需的。有趣的是，SMARCA2/4 的重定位不依赖于 γH2AX、ATM、ATR、p300/CBP 或 PARP1/2，表明它可能直接作为 DNA 损伤传感器识别 DNA 损伤。缺乏 SMARCA2/4 会延长 γH2AX、环指蛋白 8（RNF8）和乳腺癌易感基因 1（BRCA1）在 DNA 损伤部位的滞留时间，并损害 RAD51 依赖性同源重组修复。此外，SMARCA2/4 抑制剂的处理可使肿瘤细胞对 PARP 抑制剂治疗敏感。
本研究揭示了 SMARCA2/4 作为双链断裂修复的 DNA 损伤修复因子。
