Reduced Expression of GINS Complex Members Induces Hallmarks of Pre-malignancy in Primary Untransformed Human Cells

In cancer cells ablation of the GINS complex member Psf2 elicits chromosome mis-segregation yet the precise role of Psf2 in mitosis is unknown. We investigated the putative mitotic role of the GINS complex using synchronized cultures of untransformed Human Dermal Fibroblasts (HDF). Metaphase spreads from Psf1/Psf2-depleted HDF were normal and mitotic exit of Psf1/Psf2-depleted cells was only slightly delayed, suggesting no direct role for the GINS complex in mitosis of untransformed cells. Because the GINS complex is required for initiation and elongation events during DNA replication we hypothesized that the mitotic delay of Psf1/Psf2-deficient cells resulted indirectly from defective DNA synthesis during a prior S-phase. Therefore, we investigated the effects of Psf1/Psf2-depletion on DNA replication. Recruitment of Mcm7 to chromatin during G1 was unaffected by Psf1/Psf2-ablation, indicating that replication licensing does not require GINS. However, chromatin-binding of Cdc45 and PCNA and the onset of DNA synthesis were delayed in Psf1/Psf2-ablated cells. The S-phase delay of Psf1/Psf2-depleted cells was associated with hallmarks of pre-malignancy including acquisition of the DNA damage marker γH2AX, activation of Chk2, and increased Tyr 15 phosphorylation of Cdc2. Ectopic expression of catalytically-inactive Chk2 prevented Cdc2 phosphorylation and resulted in increased numbers of aberrant mitotic cells containing micronuclei, thereby recapitulating the effect of Psf2-deficiency in cancer cells. Therefore, S-phase progression of untransformed cells containing sub-optimal levels of Psf1/2 is associated with replication stress and acquisition of DNA damage. The ensuing Chk2-mediated DNA damage signalling is important for preventing mitotic defects and guarding genome integrity.