The MRE11 nuclease promotes homologous recombination not only in DNA double-strand break resection but also in post-resection in human TK6 cells

Abstract

Homologous recombination (HR) repairs double-strand breaks (DSBs) occurring in sister chromatids using the intact sisters as the repair template. HR is initiated by DSB resection, which generates 3′ single-strand DNA (ssDNA). RAD51 recombinase polymerizes on the ssDNA and undergoes strand exchange with intact sister chromatids, generating junction molecules (JMs). The separation of JMs completes HR-dependent DSB repair. Defective resolution of JMs not only leaves DSBs unrepaired but also has the broken sisters remain entangled with the intact sisters, leading to the formation of isochromatid-type breaks, where both sister chromatids are broken at the same sites, in mitotic chromosome spreads. The MRE11 nuclease plays a key role in HR, and it is generally believed that MRE11 does so by initiating DSB resection. We here showed that the loss of MRE11 reduced the efficiency of HR in human TK6 cells without affecting DSB resection, indicating a role for MRE11 in HR also at a post-resection step. MRE11-deficient TK6 cells showed proficient induction of RAD51 foci by ionizing-radiation (IR) and olaparib but significantly delayed their resolution. Although exposure of G2-phase cells to IR cleaves only one of two sister chromatids, the loss of the MRE11-nuclease activity increased the number of isochromosome-type breaks in subsequent M phase. The overexpression of GEN1 resolvase suppressed the formation of IR-induced isochromatid-type breaks in MRE11-nuclease-deficient TK6 cells. These data indicate that MRE11 plays an important role in HR by processing JMs. We propose the dual roles of MRE11 in HR at DSB resection and post-resection steps.