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. 2016 Aug 30;7(39):63887–63900. doi: 10.18632/oncotarget.11698

Figure 5. Effects of FAM122A on PP2A subunit expression and the ubiquitination of PP2A-Cα protein.

Figure 5

(A) FAM122A silencing and NC 293T cells were transiently transfected with Flag-PP2A B55α plasmid, followed by IP by anti-Flag M2 affinity gel. (B) 293T cells were transiently transfected with different concentrations of Flag tagged FAM122A or 3 μg Flag-CIP2A expressing plasmids. After transfection for 48 hours, the cell lysates were extracted and examined by western blot. (C) The protein levels of PP2A subunits from indicated 293T cells were detected by western blot. (D) The mRNA levels of PP2A subunits were examined by real-time PCR in the indicated 293T cells. (E) 293T cells were transiently cotransfected with the plasmids as indicated. After transfection for 48 hours, the cells were treated with or without MG132 at 20 μM for 4 hours. The cellular lysates were collected and immunoprecipitated with anti-Flag M2 affinity gel followed by western blot. (F) The indicated FAM122A silencing 293T cells together with NC or ectopically expressed GFP-FAM122A in siFAM#5 expressing cells were transiently transfected with Flag-PP2A-Cα plasmid respectively followed by the treatment of MG132 at 20 mM for 4 hours. IP was applied by anti-PP2A-Cα antibody and the endogenous ubiquitination of PP2A-Cα protein was analyzed by western blot.