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Fig. 5. RECQL4 is associated with regions of ssDNA and colocalizes with Rad51, but it does not colocalize with BRCA1. (A) HeLa cells were grown in BrdU-containing medium and stained for RECQL4 (red), BrdU (green) and nuclear DNA (blue). Bar, 
10 µm. (Upper panel) RecQL4 and BrdU staining in untreated cells. (Lower panel) RECQL4 colocalizes with ssDNA after treatment with 10 µM etoposide. (B) RECQL4 colocalizes with Rad51. WI38/VA13 cells were stained with anti-RECQL4 (red) and anti-Rad51 antibodies (green), and DAPI (blue). RECQL4 and Rad51 antibody-staining in untreated cells (upper panel) and after 10 µm etoposide treatment (lower panel). Bar, 10 µm. (C) RECQL4 and Rad51 form a complex in human cells. 293T cells were transiently transfected with FLAG-RECQL4, nuclear extracts derived from these cells were immunoprecipitated with the anti-FLAG antibody, and analysed by SDS-polyacrylamide gel electrophoresis. One-tenth of the same nuclear extract was used as input control (lane 1). Immunoprecipitated FLAG-RECQL4 (upper and middle panels) and Rad51 (lower panel) were detected by western blotting with the anti-FLAG, anti-RECQL4 C-t and anti-Rad51 antibodies, but not with the control IgG (lane 2). Reciprocal co-immunoprecipitation is shown in lanes 4-6; lane 4, input; lane 5, immunoprecipitation with the control IgG; lane 6, immunoprecipitation with an anti-Rad51 antibody. (D) Colocalization of RECQL4 with BRCA1 in untreated HeLa cells (upper panel) and in HeLa cells treated with 10 µM etoposide (lower panel). RECQL4 (red), BRCA1 (green), DAPI-staining shows nuclear DNA. Bar, 10 µm.
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