Functional TRPV6 channels are crucial for transepithelial Ca2+absorption

Abstract

TRPV6 is considered the primary protein responsible for transcellular Ca2+absorption. In vitro studies demonstrate that a negatively charged amino acid (D) within the putative pore region of mouse TRPV6 (position 541) is critical for Ca2+permeation of the channel. To elucidate the role of TRPV6 in transepithelial Ca2+transport in vivo, we functionally analyzed a TRPV6D541A/D541Aknockin mouse model. After weaning, mice were fed a regular (1% wt/wt) or Ca2+-deficient (0.02% wt/wt) diet and housed in metabolic cages. Blood was sampled for Ca2+measurements, and the expression of Ca2+transport proteins was analyzed in kidney and duodenum. Intestinal45Ca2+uptake was measured in vivo by an absorption assay. Challenging the mice with the Ca2+-deficient diet resulted in hypocalcemia in wild-type and TRPV6D541A/D541Amice. On a low-Ca2+diet both mouse strains displayed increased expression of intestinal TRPV6, calbindin-D9K, and renal TRPV5. TRPV6D541A/D541Amice showed significantly impaired intestinal Ca2+uptake compared with wild-type mice, and duodenal TRPV5 expression was increased in TRPV6D541A/D541Amice. On a normal diet, serum Ca2+concentrations normalized in both mouse strains. Under these conditions, intestinal Ca2+uptake was similar, and the expression levels of renal and intestinal Ca2+transport proteins were not affected. We demonstrate that TRPV6D541A/D541Amice exhibit impaired transcellular Ca2+absorption. Duodenal TRPV5 expression was increased in TRPV6D541A/D541Amice, albeit insufficient to correct for the diminished Ca2+absorption. Under normal conditions, when passive Ca2+transport is predominant, no differences between wild-type and TRPV6D541A/D541Amice were observed. Our results demonstrate a specific role for TRPV6 in transepithelial Ca2+absorption.