Figure 2
Omomyc and Myc associate to the PRMT5-MEP50 complex.
(A) Immunoprecipitation experiments. Cell extracts from U87MG/FO cells—cultured in the presence or absence of doxycycline for 24 hrs—were immunoprecipitated with either anti-Flag, anti-PRMT5 or anti-Myc antibodies. Flag-Omomyc, PRMT5, MEP50 and Myc were detected by subsequent western blot analyses. Flag-Omomyc recruited PRMT5-MEP50 and Myc proteins. Consistently, PRMT5-MEP50 recruited both Flag-Omomyc and Myc. Immunoprecipitation with the anti-Myc antibody was less efficient; however, Flag-Omomyc and PRMT5-MEP50 were still detected. (B) Immunoprecipitation from BT168/FO cell extracts after 8 and 24 hrs of doxycycicline treatment. The anti-Myc antibody was used. PRMT5-MEP50, Flag-Omomyc and Myc were found associated also in this cell line. (C) Immunoprecipitation experiments from HEK293T/FO cell extracts. Either Anti-Flag or anti-PRMT5 antibodies were used. In both the experiments, Flag-Omomyc, PRMT5-MEP50 and Myc were found associated. (D) Immunoprecipitation experiments from HEK293T cell extracts transiently co-tranfected with both PRMT5-6myc and Flag-Omomyc (FO) vectors. The Flag-Red (FR) vector was used as control. PRMT5 was found specifically associated to Flag-Omomyc. Abbreviations: TOT = total extracts; NL = nuclear extracts; IP = immunoprecipitate. All these results are representative of at least four independent experiments.
