Replication Fork Stalling in human cells: an insight into the molecular mechanisms and the impact on genome stability

DNA replication is an important process that ensures the faithful and complete duplication of the genome inside the nucleus before cell division. The replisome is the multiprotein complex that allows the replication of the genome with in particular, the DNA polymerase (Polδ) and the DNA polymerase (Polε) that are responsible for the synthesis of the lagging and leading strands, respectively. The speed of the replication fork is not uniform in eukaryotes: it can stop, pause or even arrest. Thus, Replication Fork Stalling (RFS) can make the DNA more susceptible to damage if not properly managed, threatening the stability of the genome. ChIP-Seq experiments in the laboratory have shown that RFS of POLD1, the catalytic subunit of Polδ, are located at preferential genomic sites in the human K562 cell line. In this study, we performed ChIP-qPCR experiments for various components of the replisome, like Polδ and Polε subunits and MCM proteins, to analyze RFS sites during an unperturbed S-phase or after a low aphidicolin-induced replicative stress. More particularly, we studied the Replication Fork Barrier (RFB) of the ribosomal genes and some specific genomic RFS and showed that these POLD1 RFS are as well reliable pausing sites of the replisome.