(A) BJ fibroblasts were either left untreated or treated with 5 nM actinomycin D for 8 h, followed by 50 μM of proteasomal inhibitor MG132 for 8 h. Lysates were immunoprecipitated with either control IgG or AK96 monoclonal antibody against SRSF1. Whole-cell lysates (Input) and IPs were analyzed with the indicated antibodies.
(B) BJ TT7-SRSF1 fibroblasts or empty-vector control (BJ TT7) were treated with doxycycline for 36 h and 5 nM actinomycin D for 8 h and analyzed by immunoblotting, as indicated. Values represent fold change in protein levels, relative to a loading control. Representative western blots are shown. Data are means +/− s.d. (n=3), *P<0.05, **P<0.01, *** P<0.001.
(C) Total RNA from BJ TT7-SRSF1 cells was amplified by radioactive RT-PCR and analyzed by native PAGE. Values represent fold change in mRNA levels relative to β-actin. Data are means +/− s.d. (n=3), *P<0.05, **P<0.01, *** P<0.001.
See also Figure S2.