Bursomanno Sara, Beli Petra, Khan Asif M, Minocherhomji Sheroy, Wagner Sebastian A, Bekker-Jensen Simon, Mailand Niels, Choudhary Chunaram, Hickson Ian D, Liu Ying
Center for Healthy Aging, Department of Cellular and Molecular Medicine, University of Copenhagen, Panum Institute, DK-2200 Copenhagen, Denmark.
Department of Proteomics, The Novo Nordisk Foundation Centre for Protein Research, University of Copenhagen, Panum Institute, DK-2200 Copenhagen, Denmark; Institute of Molecular Biology (IMB), Ackermannweg 4, 55128 Mainz, Germany.
DNA Repair (Amst). 2015 Jan;25:84-96. doi: 10.1016/j.dnarep.2014.10.011. Epub 2014 Nov 25.
SUMOylation is a form of post-translational modification involving covalent attachment of SUMO (Small Ubiquitin-like Modifier) polypeptides to specific lysine residues in the target protein. In human cells, there are four SUMO proteins, SUMO1-4, with SUMO2 and SUMO3 forming a closely related subfamily. SUMO2/3, in contrast to SUMO1, are predominantly involved in the cellular response to certain stresses, including heat shock. Substantial evidence from studies in yeast has shown that SUMOylation plays an important role in the regulation of DNA replication and repair. Here, we report a proteomic analysis of proteins modified by SUMO2 in response to DNA replication stress in S phase in human cells. We have identified a panel of 22 SUMO2 targets with increased SUMOylation during DNA replication stress, many of which play key functions within the DNA replication machinery and/or in the cellular response to DNA damage. Interestingly, POLD3 was found modified most significantly in response to a low dose aphidicolin treatment protocol that promotes common fragile site (CFS) breakage. POLD3 is the human ortholog of POL32 in budding yeast, and has been shown to act during break-induced recombinational repair. We have also shown that deficiency of POLD3 leads to an increase in RPA-bound ssDNA when cells are under replication stress, suggesting that POLD3 plays a role in the cellular response to DNA replication stress. Considering that DNA replication stress is a source of genome instability, and that excessive replication stress is a hallmark of pre-neoplastic and tumor cells, our characterization of SUMO2 targets during a perturbed S-phase should provide a valuable resource for future functional studies in the fields of DNA metabolism and cancer biology.
SUMO化是一种翻译后修饰形式，涉及将SUMO（小泛素样修饰物）多肽共价连接到靶蛋白中的特定赖氨酸残基上。在人类细胞中，有四种SUMO蛋白，即SUMO1 - 4，其中SUMO2和SUMO3形成一个密切相关的亚家族。与SUMO1不同，SUMO2/3主要参与细胞对某些应激的反应，包括热休克。来自酵母研究的大量证据表明，SUMO化在DNA复制和修复的调控中起重要作用。在这里，我们报告了一项蛋白质组学分析，研究人类细胞S期DNA复制应激时被SUMO2修饰的蛋白质。我们鉴定出一组22个在DNA复制应激期间SUMO化增加的SUMO2靶标，其中许多在DNA复制机制内和/或细胞对DNA损伤的反应中发挥关键功能。有趣的是，在促进常见脆性位点（CFS）断裂的低剂量阿非科林处理方案下，发现POLD3的修饰最为显著。POLD3是芽殖酵母中POL32的人类同源物，已被证明在断裂诱导的重组修复过程中起作用。我们还表明，当细胞处于复制应激时，POLD3的缺乏会导致RPA结合的单链DNA增加，这表明POLD3在细胞对DNA复制应激的反应中起作用。鉴于DNA复制应激是基因组不稳定的一个来源，并且过度的复制应激是肿瘤前细胞和肿瘤细胞的一个标志，我们对受干扰的S期期间SUMO2靶标的表征应该为DNA代谢和癌症生物学领域的未来功能研究提供有价值的资源。
