Role of the α-kinase domain in transient receptor potential melastatin 6 channel and regulation by intracellular ATP

Abstract

Transient receptor potential melastatin 6 (TRPM6) plays an essential role in epithelial Mg2+ transport. TRPM6 and its closest homologue, TRPM7, both combine a cation channel with an α-kinase domain. However, the role of this α-kinase domain in TRPM6 channel activity remains elusive. The aim of this study was to investigate the regulation of TRPM6 channel activity by intracellular ATP and the involvement of its α-kinase domain. We demonstrated that intracellular Na- and Mg-ATP decreased the TRPM6 current in HEK293 cells heterogeneously expressing the channel, whereas Na-CTP or Na-GTP had no effect on channel activity. Whole cell recordings in TRPM6-expressing HEK293 cells showed that deletion of the α-kinase domain prevented the inhibitory effect of intracellular ATP without abrogating channel activity. Mutation of the conserved putative ATP-binding motif GXG(A)XXG (G1955D) in the α-kinase domain of TRPM6 inhibited the ATP action, whereas this effect remained preserved in the TRPM6 phosphotransferase-deficient mutant K1804R. Mutation of the TRPM6 autophosphorylation site, Thr1851, into either an alanine or an aspartate, resulted in functional channels that could still be inhibited by ATP. In conclusion, intracellular ATP regulates TRPM6 channel activity via its α-kinase domain independently of α-kinase activity. © 2008 by The American Society for Biochemistry and Molecular Biology, Inc.