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. 2013 Mar 14;288(16):11348–11357. doi: 10.1074/jbc.M112.429134

FIGURE 3.

FIGURE 3.

ERα is poly(ADP-ribosy)lated by PARP1. A, VSMCs were transfected with PARP1 siRNA (50 nm) or unrelated siRNA (50 nm) for 48 h. The poly(ADP-ribosyl)ation levels in VSMCs were determined by Western blotting with anti-PAR Ab. GAPDH was used for standardization. B, in a cell-free system, recombinant ERα protein was incubated with vehicles, PARP1/NAD+/active DNA, or PARP1/NAD+/active DNA/3AB, respectively. Poly(ADP-ribosyl)ation levels of recombinant ERα were determined by Western blotting with anti-PAR Ab. Western blotting with anti-ERα Ab showed the loading control. The results are from three independent experiments. C, VSMCs were treated with estradiol (E, 10−8, 10−7, 10−6 m, 24 h), or transfected with PARP1 siRNA (50 nm, 48 h) or unrelated siRNA (50 nm, 48 h). Immunoprecipitation (IP) of ERα from VSMCs treated as indicated, followed by Western blot assay using anti-PAR antibody. D, VSMCs were treated with estradiol (E, 10−8, 10−7, 10−6 m, 24 h). PARP activity in VSMCs was assayed as described under “Experimental Procedures.” The results are from six independent experiments. Data are expressed as the mean ± S.E. #, p < 0.05 is for comparison with control group.