Institute of Molecular Biology, Academia Sinica, Taipei 115, Taiwan.
Laboratory of Genome-Chromosome Functions, Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Japan.
Nucleic Acids Res. 2020 Sep 4;48(15):8474-8489. doi: 10.1093/nar/gkaa587.
Highly toxic DNA double-strand breaks (DSBs) readily trigger the DNA damage response (DDR) in cells, which delays cell cycle progression to ensure proper DSB repair. In Saccharomyces cerevisiae, mitotic S phase (20-30 min) is lengthened upon DNA damage. During meiosis, Spo11-induced DSB onset and repair lasts up to 5 h. We report that the NH2-terminal domain (NTD; residues 1-66) of Rad51 has dual functions for repairing DSBs during vegetative growth and meiosis. Firstly, Rad51-NTD exhibits autonomous expression-enhancing activity for high-level production of native Rad51 and when fused to exogenous β-galactosidase in vivo. Secondly, Rad51-NTD is an S/T-Q cluster domain (SCD) harboring three putative Mec1/Tel1 target sites. Mec1/Tel1-dependent phosphorylation antagonizes the proteasomal degradation pathway, increasing the half-life of Rad51 from ∼30 min to ≥180 min. Our results evidence a direct link between homologous recombination and DDR modulated by Rad51 homeostasis.
高度有毒的 DNA 双链断裂 (DSB) 很容易在细胞中引发 DNA 损伤反应 (DDR)，这会延迟细胞周期进程以确保正确的 DSB 修复。在酿酒酵母中，有丝分裂 S 期（20-30 分钟）在 DNA 损伤后延长。在减数分裂过程中，Spo11 诱导的 DSB 起始和修复持续长达 5 小时。我们报告说，Rad51 的 NH2 末端结构域（NTD；残基 1-66）在营养生长和减数分裂过程中具有修复 DSB 的双重功能。首先，Rad51-NTD 表现出自发表达增强活性，可高水平生产天然 Rad51，并且在体内与外源β-半乳糖苷酶融合时也是如此。其次，Rad51-NTD 是一个 S/T-Q 簇结构域 (SCD)，含有三个推定的 Mek1/Tel1 靶位。Mek1/Tel1 依赖性磷酸化拮抗蛋白酶体降解途径，使 Rad51 的半衰期从约 30 分钟增加到≥180 分钟。我们的结果证明了同源重组和 DDR 之间的直接联系，由 Rad51 动态平衡调节。
