Cloning and characterization of PIMT, a protein with a methyltransferase domain, which interacts with and enhances nuclear receptor coactivator PRIP function

The nuclear receptor coactivators participate in the transcriptional activa tion of specific genes by nuclear receptors. in this study, we report the i solation of a nuclear receptor coactivator-interacting protein from a human liver cDNA library by using the coactivator peroxisome proliferator-activa ted receptor-interacting protein (PRIP) (ASC2/AIB3/RAP250/NRC/TRBP) as bait in a yeast two-hybrid screen. Human PRIP-interacting protein cDNA has an O RF of 2,556 nucleotides, encodes a protein with 852 amino acids, and contai ns a 9-aa VVDAFCGVG methyltransferase motif I and an invariant GXXGXXI segm ent found in K-homology motifs of many RNA-binding proteins. The gene encod ing this protein, designated PRIP-interacting protein with methyltransferas e domain (PIMT), is localized on chromosome 8q11 and spans more than 40 kb. PIMT mRNA is ubiquitously expressed, with a high level of expression in he art, skeletal muscle, kidney, liver, and placenta. Using the immunofluoresc ence localization method, we found that PIMT and PRIP proteins appear coloc alized in the nucleus. PIMT strongly interacts with PRIP under in vitro and in vivo conditions, and the PIMT-binding site on PRIP is in the region enc ompassing amino acids 773-927. PIMT binds S-adenosyl-L-methionine, the meth yl donor for methyltransfer reaction, and it also binds RNA, suggesting tha t it is a putative RNA methyltransferase. PIMT enhances the transcriptional activity of peroxisome proliferator-activated receptor gamma and retinoid- X-receptor alpha, which is further stimulated by coexpression of PRIP, impl ying that PIMT is a component of nuclear receptor signal transduction appar atus acting through PRIP. Definitive identification of the specific substra te of PIMT and the role of this RNA-binding protein in transcriptional regu lation remain to be determined.