Seong Changhyun, Sehorn Michael G, Plate Iben, Shi Idina, Song Binwei, Chi Peter, Mortensen Uffe, Sung Patrick, Krejci Lumir
Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
J Biol Chem. 2008 May 2;283(18):12166-74. doi: 10.1074/jbc.M800763200. Epub 2008 Feb 29.
A helical filament of Rad51 on single-strand DNA (ssDNA), called the presynaptic filament, catalyzes DNA joint formation during homologous recombination. Rad52 facilitates presynaptic filament assembly, and this recombination mediator activity is thought to rely on the interactions of Rad52 with Rad51, the ssDNA-binding protein RPA, and ssDNA. The N-terminal region of Rad52, which has DNA binding activity and an oligomeric structure, is thought to be crucial for mediator activity and recombination. Unexpectedly, we find that the C-terminal region of Rad52 also harbors a DNA binding function. Importantly, the Rad52 C-terminal portion alone can promote Rad51 presynaptic filament assembly. The middle portion of Rad52 associates with DNA-bound RPA and contributes to the recombination mediator activity. Accordingly, expression of a protein species that harbors the middle and C-terminal regions of Rad52 in the rad52 Delta327 background enhances the association of Rad51 protein with a HO-made DNA double-strand break and partially complements the methylmethane sulfonate sensitivity of the mutant cells. Our results provide a mechanistic framework for rationalizing the multi-faceted role of Rad52 in recombination and DNA repair.
单链DNA（ssDNA）上由Rad51组成的螺旋状细丝，即突触前细丝，在同源重组过程中催化DNA连接的形成。Rad52促进突触前细丝的组装，并且这种重组介导活性被认为依赖于Rad52与Rad51、ssDNA结合蛋白RPA以及ssDNA之间的相互作用。Rad52的N端区域具有DNA结合活性和寡聚结构，被认为对介导活性和重组至关重要。出乎意料的是，我们发现Rad52的C端区域也具有DNA结合功能。重要的是，单独的Rad52 C端部分就能促进Rad51突触前细丝的组装。Rad52的中间部分与结合DNA的RPA相互作用，并对重组介导活性有贡献。因此，在rad52 Delta327背景中表达含有Rad52中间和C端区域的蛋白质物种，可增强Rad51蛋白与HO产生的DNA双链断裂的结合，并部分弥补突变细胞对甲基磺酸甲酯的敏感性。我们的结果为阐明Rad52在重组和DNA修复中的多方面作用提供了一个机制框架。
