Methylation of GPS2 by PRMT6 prevents its proteasomal degradation.
A, in vitro methylation assay showing GPS2 direct methylation by recombinant PRMT6. WB, Western blot. B, Western blot showing that expression of HA-GPS2 in transiently transfected 293T cells is significantly impaired upon down-regulation of PRMT6 but not of PRMT1. Blotting for PRMT1 and PRMT6 validated the efficiency of siRNAs, and blotting for tubulin provided a loading control. C, the same experimental approach was used to confirm that HA-GPS2 expression in the absence of PRMT6 can be rescued by treatment with the proteasome inhibitor MG132. Blotting for TBL1 was used as a loading control in this case. D, a significant decrease in GPS2 protein level was observed in 293T nuclear extracts upon PRMT6 down-regulation via specific siRNA, as validated by blotting for PRMT6. E, quantitative RT-PCR showing that down-regulation of PRMT6 in 293T cells by siRNA does not change the mRNA expression of GPS2. Data are mean ± S.D. **, p < 0.05.