Figure 2.

XPA-associated DSBs form at replication forks in HGPS. A) HGPS cells treated with or without CPT were subjected to the modified ChIP assay with pulldown by the anti-γ-H2AX antibody. The immunoprecipitated chromatin was analyzed for PCNA and MCM7 Co-IP by Western blot analysis. PCNA was largely absent at the replication forks with endogenous progerin-induced strand breaks, but was present in chromatin at CPT-induced strand breaks. MCM7 is a replication fork marker. B) HGPS cells were transfected with XPA siRNA, then mock or CPT treated, followed by analysis using modified ChIP assay with anti-γ-H2AX antibody. The successful knockdown of XPA by siRNA significantly restores the binding of PCNA and Polδ to the replication forks in the progeroid cells.