FIGURE 5.

Processing of telomere replication is hampered by the absence of BRCA2. A, HeLa cells were transfected with siRNA for GFP (siGFP) or BRCA2 (siBRCA2). The cells were then pulsed with BrdU for 30 min and chased for 3 h and 30 min. This cycle was repeated six times. Lysates were incubated with anti-TRF1 or -TRF2 antibodies and subjected to telomere-ChIP. The blot was immunoblotted with anti-BrdU antibodies to assess the amount of replicated telomeric DNAs. ChIP assays were also performed with Alu or telomere probes. The results from two different sets of experiments are marked below the left panel. The values in percentage depict the ratio of BrdU incorporation in BRCA2-depleted cells (siBRCA2) compared with control (siGFP). B, flow cytometry analysis of cell cycle progression with or without BRCA2 depletion in HeLa cells. The percentage of cells in S phase is boxed and marked (right panel). Simultaneously, lysates were prepared for Western blot (WB) to assess the efficiency of siRNA against BRCA2. Same blot was probed with anti-lamin A/C for normalization (left panel).