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Cell cycle-dependent processing of DNA lesions controls localization of Rad9 to sites of genotoxic stress. | LitMetric

The Rad9/Rad1/Hus1 complex functions to facilitate the ATR-mediated phosphorylation of several substrates that control the checkpoint arrest induced by DNA damage. Here we show that in response to genotoxic stress induced by different types of damaging agents, Rad9 rapidly relocalized to sites of single stranded DNA, as visualized by discrete nuclear foci that co-localize with RPA. UV light-induced Rad9 foci also colocalized with TopBP1 and gamma-H2AX. Interestingly, Rad9 foci were predominately formed in G(1) and S phase after UV light, while treatment of cells with ionizing radiation (IR) resulted in accumulation of Rad9 into foci in S and G(2). Photobleaching experiments in living cells revealed that the Rad9 protein is highly mobile in undamaged cells. However, genotoxic stress induced the immobilization of a large proportion of the protein. The proportion of Rad9 immobilization was larger in S phase and the accumulation to sites of locally damaged areas induced by UV-laser irradiation was faster during DNA replication. Inactivation of nucleotide excision repair by knock down of XPA and XPC resulted in a decrease of G(1) phase cells that displayed Rad9 foci in response to UV light, whereas IR-induced Rad9 foci were not affected. In contrast, downregulation of CtIP, which promotes DSB resection, abrogated the IR-induced Rad9 foci. These findings show that due to processing of DNA lesions into a common intermediate, which occurs in a cell cycle-dependent manner, Rad9 is able to respond to different types of genotoxic stress.

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http://dx.doi.org/10.4161/cc.8.11.8721DOI Listing

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