Magalhaes Yuli T, Silva Gisele E T, Osaki Juliana H, Rocha Clarissa R R, Forti Fabio L
Biomolecular Systems Signaling Laboratory, Department of Biochemistry, Institute of Chemistry, University of São Paulo, São Paulo, Brazil.
DNA Repair Laboratory, Department of Microbiology, Biomedical Sciences Institute, University of São Paulo, São Paulo, Brazil.
Front Cell Dev Biol. 2020 Aug 19;8:816. doi: 10.3389/fcell.2020.00816. eCollection 2020.
Typical Rho GTPases include the enzymes RhoA, Rac1, and Cdc42 that act as molecular switches to regulate essential cellular processes in eukaryotic cells such as actomyosin dynamics, cell cycle, adhesion, death and differentiation. Recently, it has been shown that different conditions modulate the activity of these enzymes, but their functions still need to be better understood. Here we examine the interplay between RhoA and the NER (Nucleotide Excision Repair) pathway in human cells exposed to UVA, UVB or UVC radiation. The results show high levels and accumulation of UV-induced DNA lesions (strand breaks and cyclobutane pyrimidine dimers, CPDs) in different cells with RhoA loss of function (), either by stable overexpression of negative dominant RhoA (RhoA-N19 mutant), by inhibition with C3 toxin or by transient silencing with siRNA. Cells under RhoA showed reduced levels of γH2AX, p-Chk1 (Ser345) and p-p53 (Ser15) that reflected causally in their accumulation in G1/S phases, in low survival rates and in reduced cell proliferation, also in accordance with the energy of applied UV light. Even NER-deficient cells (XPA, XPC) or DNA translesion synthesis (TLS)-deficient cells (XPV) showed substantial hypersensitivity to UV effects when previously submitted to RhoA . In contrast, analyses of apoptosis, necrosis, autophagy and senescence revealed that all cells displaying normal levels of active RhoA (RhoA-GTP) are more resistant to UV-promoted cell death. This work reaffirms the role of RhoA protein signaling in protecting cells from damage caused by UV radiation and demonstrates relevant communicating mechanisms between actin cytoskeleton and genomic stability.
典型的Rho GTP酶包括RhoA、Rac1和Cdc42等酶，它们作为分子开关，调节真核细胞中的基本细胞过程，如肌动球蛋白动力学、细胞周期、黏附、死亡和分化。最近的研究表明，不同的条件会调节这些酶的活性，但其功能仍有待进一步了解。在这里，我们研究了在暴露于UVA、UVB或UVC辐射的人类细胞中，RhoA与核苷酸切除修复（NER）途径之间的相互作用。结果显示，在不同的RhoA功能丧失的细胞中，紫外线诱导的DNA损伤（链断裂和环丁烷嘧啶二聚体，CPD）水平较高且有积累，这些细胞通过稳定过表达负显性RhoA（RhoA-N19突变体）、用C3毒素抑制或用小干扰RNA瞬时沉默来实现RhoA功能丧失。RhoA功能丧失的细胞中，γH2AX、p-Chk1（Ser345）和p-p53（Ser15）水平降低，这因果性地反映在它们在G1/S期的积累、低存活率和细胞增殖减少上，这也与所施加紫外线的能量一致。甚至NER缺陷细胞（XPA、XPC）或DNA跨损伤合成（TLS）缺陷细胞（XPV）在先前经历RhoA功能丧失后，对紫外线效应也表现出明显的超敏性。相反，对凋亡、坏死、自噬和衰老的分析表明，所有显示正常活性RhoA（RhoA-GTP）水平的细胞对紫外线促进的细胞死亡更具抗性。这项工作重申了RhoA蛋白信号传导在保护细胞免受紫外线辐射损伤中的作用，并证明了肌动蛋白细胞骨架与基因组稳定性之间的相关通讯机制。
