Fig. 7: Degradation of stressed forks requires PFN1 and SNF2H.

a Ratios of CIdU/IdU tract lengths in control or PFN1-overexpressing HeLa cells transfected with indicated siRNAs. b–e Ratios of CIdU/IdU tract lengths in control or PFN1 knockdown MCF-10A cells transfected with the indicated siRNAs. f Ratios of CIdU/IdU tract lengths in control or SNF2H knockdown MCF-10A cells. g, h Ratios of CIdU/IdU tract lengths in control or PFN1 knockdown or overexpression MCF-10A cells with or without SNF2H knockdown. i, j Ratios of CIdU/IdU in control or SNF2H knockdown MCF-10A cells transfected with control or gene-specific siRNAs. Cells in a–j were all dual-labeled with IdU-CIdU and subsequently treated with 4 mM HU for 2 h. Statistical significance was determined by Kruskal–Wallis test with Dunnett’s multiple comparisons. ****p < 0.0001; ns, not significant. In a–j, at least 300 fibers were analyzed per condition. All results were confirmed by at least two independent experiments. k Working model of the context-dependent regulation of DNA replication forks by PFN1 via its interactions with SNF2H and BOD1L. Under normal conditions, we hypothesize that the physical and functional interaction of PFN1 with SNF2H increases parental chromatin relaxation needed for both DNA replication origin firing and replication fork progression. Under stressed conditions, we hypothesize that PFN1/SNF2H remodels nucleosomes on nascent chromatin to enable general fork reversal mediated by different DNA translocases and helicases. In addition, direct binding and inhibition of BOD1L by PFN1 causes destabilization of a subset of reversed forks which are protected by BOD1L-dependent RAD51 nucleofilaments.