Human SMC5/6 complex promotes sister chromatid homologous recombination by recruiting the SMC1/3 cohesin complex to double-strand breaks

Abstract

The structural maintenance of chromosomes (SMC) family of proteins has been implicated in the repair of DNA double-strand breaks (DSBs) by homologous recombination (HR). The SMC1/3 cohesin complex is thought to promote HR by maintaining the close proximity of sister chromatids at DSBs. The SMC5/6 complex is also required for DNA repair, but the mechanism by which it accomplishes this is unclear. Here, we show that RNAi-mediated knockdown of the SMC5/6 complex components in human cells increases the efficiency of gene targeting due to a specific requirement for hSMC5/6 in sister chromatid HR. Knockdown of the hSMC5/6 complex decreases sister chromatid HR, but does not reduce nonhomologous endjoining (NHEJ) or intra-chromatid, homologue, or extrachromosomal HR. The hSMC5/6 complex is itself recruited to nuclease-induced DSBs and is required for the recruitment of cohesin to DSBs. Our results establish a mechanism by which the hSMC5/6 complex promotes DNA repair and suggest a novel strategy to improve the efficiency of gene targeting in mammalian somatic cells.

Smc5–Smc6 complex suppresses gross chromosomal rearrangements mediated by break-induced replications

“…For example, the cohesion defect impairs sister chromatid recombination [31,66], increasing intra-chromatid recombination in the tandem array of ribosomal repeats [36]. Recent work even suggested that Smc5-Smc6 would be required for the efficient loading of cohesin to DSB sites [67,68]. Smc5-Smc6 could restrain the initiation of strand invasions between different chromosomes that lead to BIR-mediated translocation events.…”

Nuclear organization in genome stability: SUMO connections

“…Mutations in genes involved in the SUMO pathway, such as Ubc9, Mms21, Ulp1, Slx5 and Slx8, render cells sensitive to DNA damage, and many proteins involved in the maintenance of genome stability have been shown to become sumoylated during the damage response [23][24][25][26][27][28]. Specifically, it has been shown that the SUMO pathway plays an important role in homologous recombination and the resumption of stalled/collapsed replication forks [27,[36][37][38].…”

“…This complex is specifically required for recombination between sister chromatids in both yeast [37] and man [36], and is recruited to DSBs in G2/M cells [37,42]. Moreover, it contributes to the maintenance of telomeric repeats by recombination in telomerasedeficient cells [21].…”

“…The resolution of the joint molecule completes this error-free repair event. The Smc5/Smc6 complex, which binds a SUMO E3 ligase Mms21, is required for sister chromatid exchange in both yeast [37] and man [36]. (B) When DNA replication forks encounter a damaged base (shown as a circle), it often leads to the collapse of replication forks, resulting in a one-end DSB.…”

APRIN is a cell cycle specific BRCA2-interacting protein required for genome integrity and a predictor of outcome after chemotherapy in breast cancer

“…Since APRIN appeared to modulate HR capacity, we investigated whether APRIN was localised at the site of a DSB. To assess this, we designed a chromatin IP (ChIP) assay based upon the HR reporter described in Figure 4C and similar to that previously described by Potts and colleagues (Potts et al, 2006). Cells harbouring a genomic DR-GFP reporter were transfected with an I-SceI expression construct that caused a DSB within the reporter construct.…”