As part of our studies on the membrane-initiated actions of 1,25(OH)₂D_3, and its localization in caveolae membrane fractions, we used a VDR-KO mouse model to study the binding of [³H]-1,25(OH)₂D₃ in the presumed absence of the VDR. In this mouse model, known as the Tokyo strain, the second exon of the VDR gene, which encodes the first of the two zinc fingers responsible for DNA binding, was removed and the resulting animals have been considered to be VDR-null mice. To our surprise, several tissues in these KO mice showed significant (5 to 50% of that seen in WT animals). specific binding of [³H]-1,25(OH)₂D₃ in nuclear and caveolae membrane fractions The K_D's of this binding in samples from VDR-KO and wild-type mice were indistinguishable. RT-PCR analysis of intestinal mRNA from the VDR-KO animals revealed an mRNA that lacks exon 2 but contains exons 3-9 plus two 5'-untranslated exons. Western analysis of intestinal extracts from VDR-KO mice showed a protein of a size consistent with the use of Met52 as the translational start site. Transfection of a plasmid construct containing the sequence encoding the human analog of this truncated form of the receptor, VDR(52-C), into Cos-1 cells showed that this truncated form of the receptor retains full [³H]-1,25(OH)₂D₃ binding ability. This same construct was inactive in transactivation assays using the osteocalcin promoter in CV1 cells. Thus, we have determined that this widely used strain of the VDR-KO mouse can express a form of the VDR that can bind ligand but not activate gene transcription.