VDR mutant M1T protein has diminished protein‐protein interactions

Abstract

The Vitamin D3 Receptor (VDR) is a steroid hormone receptor protein that forms homodimers and heterodimers. The Retinoid X Receptor‐alpha receptor (RXRa) protein is a heterodimerization binding partner for VDR. A start codon polymorphism in the VDR gene, known as M1T VDR, has been reported. This VDR gene variant has been associated with increased risk for some human cancers. The mutant M1T VDR mRNA initiates protein translation at an internal Met codon and produces M1T VDR protein lacking the first three amino acids present in full length VDR protein. The goal of this work was to detect any differences in protein‐protein interactions formed by M1T VDR with VDR, M1T VDR or RXRa. We have constructed six mammalian expression vectors that produces either untagged VDR, M1T VDR, or RXRa, as well as, N‐terminally epitope‐tagged myc‐VDR, myc‐M1T VDR or myc‐ RXRa. The experimental approached employed in this study involves transient co‐transfection of H1299 cells with combinations of untagged and myc‐tagged expression vectors, followed by co‐immunoprecipitation (Co‐IP) assays and Western blotting for VDR, M1T VDR or RXRa. Our data are consistent with at least a 50% reduction in the ability of M1T VDR to form homodimers and heterodimers. These data suggest a potential molecular etiology for increased cancer risks associated with the M1T VDR mutant gene.