Autoinhibition of TRPV6 Channel by Intramolecular Interactions

Transient receptor potential vanilloid 6 (TRPV6), a calcium-selective channel possessing six transmembrane domains (S1-S6) and intracellular N-/C-termini, plays crucial roles in calcium absorption in epithelia and bone and is involved in human diseases including vitamin-D deficiency, osteoporosis and cancer. The TRPV6 function and regulation remain poorly understood. Here by electrophysiology and Xenopus oocyte expression we found that Arg510 in the S4-S5 linker and Trp633 in the TRP helix are functionally critical. By co-immunoprecipitation and in vitro binding we found that the Arg510:Trp633 pair mediates binding between the linker (L) and C-terminal TRP helix (C) and that disruption of the linker/TRP helix (L/C) interaction substantially increases the channel activity. We also found that Trp361 in the N-terminal pre-S1 helix (N) and Ile637 mediate the pre-S1/TRP helix (N/C) binding which is functionally critical as well. Calcium imaging in human embryonic kidney cells further supported functional importance of Trp361 and Trp633. Besides, disruption of either interaction by blocking peptides activated TRPV6. The L/C interaction was required for the N/C interaction but not reversely. Together, we showed that the TRPV6 intramolecular S4-S5 linker to TRP helix and pre-S1 helix to TRP helix interactions, mediated by Arg510:Trp633 and Trp361:Ile637 bonding, respectively, are autoinhibitory and are required for maintaining TRPV6 at basal states. This study reveals a regulatory mechanism of TRPV6 activation-autoinhibition, which would help elucidating the corresponding mechanisms in other TRP channels. Supported by AIGSS (to RC), NSERC and KFoC (to XZC).