Title:
A hidden DNA synthesis in vertebrate cells

Masato Kanemaki
Center for Frontier Research, National Institute of Genetics, Yata 1111, Mishima, Shizuoka 411-8540, Japan

Abstract:
Recombination repair between sister chromatid DNAs plays a crucial role when replication forks encounter to a block (such as inter-strand crosslink, ICL) that impedes progression of the replicative Mcm2-7 helicase or when double-strand breaks are introduced to post-replicative DNA.  Rad51 promotes strand invasion of ssDNA having a 3’ end for initiation of DNA synthesis in order to copy the intact sequence on the sister chromatid.  Mechanisms leading to the Rad51-dependent strand invasion are relatively well understood.  However, little is known how DNA synthesis, which presumably requires both DNA polymerase and helicase, is carried out after strand invasion in vertebrate cells.
We recently reported that Mcm8 and Mcm9, both of which evolved from the same ancestor of Mcm2-7, form a stable complex that functions in recombination repair in chicken DT40 cells (Nishimura et al, Mol Cell, 2012).  Why are the Mcm-related proteins involved in recombination repair, but not in normal DNA replication?  This might be because the Mcm8-9 complex plays a unique role in a crosstalk between DNA replication and recombination repair.
By employing the CRISPR-based gene editing technology, we generated human cells lacking theMCM8 or MCM9 gene.  We found that these human KO cells showed hypersensitivity to ICL-inducing agents, but not to ionizing radiation (IR), suggesting that a specific recombination involving Mcm8-9 might be required for ICL repair.  In both human and chicken DT40 cells, our current results support the idea that the Mcm8-9 complex functions after Rad51 loading at damaged sites.  Furthermore, recent genetic and biochemical data strongly suggest that the Mcm8-9 complex promotes DNA strand synthesis as a DNA helicase.  By presenting a model of the role for the Mcm8-9 complex, the relationship between DNA replication and recombination-dependent DNA strand synthesis will be discussed.