MCM8IP activates the MCM8-9 helicase to promote DNA synthesis and homologous recombination upon DNA damage

Abstract

Homologous recombination (HR) mediates the error-free repair of DNA double-strand breaks to maintain genomic stability. Here we characterize C17orf53/MCM8IP, an OB-fold containing protein that binds ssDNA, as a DNA repair factor involved in HR. MCM8IPdeficient cells exhibit HR defects, especially in long-tract gene conversion, occurring downstream of RAD51 loading, consistent with a role for MCM8IP in HR-dependent DNA synthesis. Moreover, loss of MCM8IP confers cellular sensitivity to crosslinking agents and PARP inhibition. Importantly, we report that MCM8IP directly associates with MCM8-9, a helicase complex mutated in primary ovarian insufficiency, and RPA1. We additionally show that the interactions of MCM8IP with MCM8-9 and RPA facilitate HR and promote replication fork progression and cellular viability in response to treatment with crosslinking agents. Mechanistically, MCM8IP stimulates the helicase activity of MCM8-9. Collectively, our work identifies MCM8IP as a key regulator of MCM8-9-dependent DNA synthesis during DNA recombination and replication.

Cockayne Syndrome B protein selectively interacts and resolves intermolecular DNA G-quadruplex structures

“…1e). Indeed, the gel quantification revealed a statistically significant increase (p=0.02 using a two-tailed Student's t-test 24 , see Supplementary Fig. S1a and Supplementary Table S2 for all the plotted data) of the percentage of ds product formed after incubation (0.5 to 40 min) with CSB (24.6%) compared to the control without the protein (8%), but the G4 resolution remains partial and, most importantly, does not increase with increasing incubation times (Fig.…”

Loss of HMCES is synthetic lethal with APOBEC activity in cancer cells

“…We prioritized genes by an overall APOBEC essentiality score: average log2 fold-change (LFC) over six measurements: three time points for the A549 TP53-/- cell line (T9, T12 and T15) and three for the LXF-289 cell line (T5, T10 and T15). The top five hits by this score were the genes coding for the AP-site protecting protein HMCES[42], the RAD9A cell cycle checkpoint control protein, the MCM8 component of the MCM8-MCM9-HROB complex[37,38,53], ATXN7L3, a component of the SAGA chromatin modifying complex[54,55], and HGC6.3, an uncharacterized protein. For four of the five genes, the individual gRNAs, four per gene, consistently sensitized cells to A3A expression (Fig 2B and S3 Fig).…”

“…Furthermore, nucleotide excision repair (NER) and DNA mismatch repair (MMR) were enriched in both cell lines, as well as the regulation of the cell cycle (Fig 2D and S3 Table). In LXF-289 cells, the non-homologous end-joining (NHEJ) and Fanconi anemia pathways were strongly represented among the top hits enriched, as well as MCM8, MCM9 and HROB, genes that have previously been implicated in HR (Fig 2D and S3 Table)[37,38,53]. Further enriched pathways related to DNA repair included error-prone TLS and telomere maintenance in LXF-289 cells.…”

Loss of the abasic site sensor HMCES is synthetic lethal with the activity of the APOBEC3A cytosine deaminase in cancer cells

“…We prioritized genes by an overall APOBEC essentiality score: average log 2 fold change (LFC) over 6 measurements: 3 time points for the A549 TP53−/− cell line (T9, T12, and T15) and 3 for the LXF-289 cell line (T5, T10, and T15). The top 5 hits by this score were the genes coding for the AP site protecting protein HMCES [42], the RAD9A cell cycle checkpoint control protein, the MCM8 component of the MCM8-MCM9-HROB complex [37,38,55], ATXN7L3, a component of the SAGA chromatin modifying complex [56,57] Next highest-ranking hits included the UBA6 ubiquitin-activating enzyme and a further 5 genes that were all related to DNA repair, DNA replication, or cell cycle control (DDX11, MCM9, CDC23, MAD2L2 (also known as REV7), and HROB (also known as C17ORF53 or MCM8IP); S1 Table ).…”

“…We performed a CRISPR/Cas-9-based genome-wide screen for genes required to tolerate A3A-mediated DNA damage in a panel of cell lines from non-small cell lung cancer (NSCLC), where APOBEC activity has been shown to play an important role in tumor evolution [3,4,[34][35][36]. Among other hits, we identified factors involved in multiple DSB repair pathways, including RAD9A, a component of the 9-1-1 alternative clamp loader and the recently characterized MCM8-MCM9-HROB complex [37][38][39][40]. Crucially, we found that different genetic backgrounds are consistently and strongly dependent on the gene encoding HMCES (5hmC binding, embryonic stem cell-specific protein) for cell viability [8,15] under APOBEC stress, but not otherwise.…”