FIGURE 4.

Comparison of DNA end resection activities of WRN-DNA2 and BLM-DNA2. A, comparison of helicase activities of WRN and BLM. Reactions contained 1 nm ³²P-labeled forked DNA duplex (inset) and different concentrations of WRN or BLM. Reactions were incubated at 37 °C for 30 min, and reaction products were quantified as described under “Experimental Procedures.” Data are mean ± S.D. (n = 3). B, time course of resection of 3′-tailed DNA substrate catalyzed by WRN-DNA2 and BLM-DNA2, respectively. Reactions contained 2 nm DNA, 350 nm RPA, 8 nm DNA2, and 10 nm WRN/BLM. Reaction aliquots withdrawn at the indicated time points were subjected to electrophoresis on a 1% agarose gel after hybridization of radiolabeled probes complementary to 3′-terminated strand at the indicated positions. Radiolabeled DNA species were visualized by phosphorimaging. C, quantification of the reactions in B. Relative concentration of resection products generated at each time point was calculated as a percentage of the product generated by 20 nm EXO1 after 2 min. Data are mean ± S.D. (n = 3). D, processing of 3′-tailed (26 nt) and blunt-ended DNA substrates in reactions with indicated composition. Reactions were carried out at 37 °C for 60 min and contained 2 nm DNA, 350 nm RPA, and, where indicated, 8 nm DNA2, 20 nm WRN, and 20 nm BLM. Reaction products were analyzed as in Fig. 1C. Lane 1, heat-denatured substrate; lane 14, 3′-tailed substrate incubated with 20 nm EXO1 for 2 min (R1); lane 15, blunt-ended substrate incubated with 20 nm EXO1 for 2 min (R2).