Department of Molecular and Cellular Biology, University of California Davis, Davis, California, United States of America.
Biochemistry, Molecular, Cellular and Developmental Biology Graduate Group, University of California Davis, Davis, California, United States of America.
PLoS Genet. 2023 Jan 30;19(1):e1010457. doi: 10.1371/journal.pgen.1010457. eCollection 2023 Jan.
The tumor suppressor BRCA1-BARD1 complex regulates many cellular processes; of critical importance to its tumor suppressor function is its role in genome integrity. Although RING E3 ubiquitin ligase activity is the only known enzymatic activity of the complex, the in vivo requirement for BRCA1-BARD1 E3 ubiquitin ligase activity has been controversial. Here we probe the role of BRCA1-BARD1 E3 ubiquitin ligase activity in vivo using C. elegans. Genetic, cell biological, and biochemical analyses of mutants defective for E3 ligase activity suggest there is both E3 ligase-dependent and independent functions of the complex in the context of DNA damage repair and meiosis. We show that E3 ligase activity is important for nuclear accumulation of the complex and specifically to concentrate at meiotic recombination sites but not at DNA damage sites in proliferating germ cells. While BRCA1 alone is capable of monoubiquitylation, BARD1 is required with BRCA1 to promote polyubiquitylation. We find that the requirement for E3 ligase activity and BARD1 in DNA damage signaling and repair can be partially alleviated by driving the nuclear accumulation and self-association of BRCA1. Our data suggest that in addition to E3 ligase activity, BRCA1 may serve a structural role for DNA damage signaling and repair while BARD1 plays an accessory role to enhance BRCA1 function.
肿瘤抑制因子 BRCA1-BARD1 复合物调节许多细胞过程；对其肿瘤抑制功能至关重要的是其在基因组完整性中的作用。尽管 RING E3 泛素连接酶活性是该复合物唯一已知的酶活性，但 BRCA1-BARD1 E3 泛素连接酶活性在体内的需求一直存在争议。在这里，我们使用秀丽隐杆线虫来探究 BRCA1-BARD1 E3 泛素连接酶活性在体内的作用。对 E3 连接酶活性缺陷突变体的遗传、细胞生物学和生化分析表明，在 DNA 损伤修复和减数分裂过程中，该复合物既有 E3 连接酶依赖性，也有非依赖性功能。我们表明，E3 连接酶活性对于复合物的核积累很重要，特别是对于在有丝分裂重组位点的浓缩，但对于增殖性生殖细胞中的 DNA 损伤位点则不然。虽然 BRCA1 本身能够进行单泛素化，但 BARD1 与 BRCA1 一起对于促进多泛素化是必需的。我们发现，E3 连接酶活性和 BARD1 在 DNA 损伤信号转导和修复中的需求可以通过驱动 BRCA1 的核积累和自组装来部分缓解。我们的数据表明，除了 E3 连接酶活性之外，BRCA1 可能在 DNA 损伤信号转导和修复中发挥结构作用，而 BARD1 则起到增强 BRCA1 功能的辅助作用。
