Molecular details of ruthenium red pore block in TRPV channels

Transient receptor potential vanilloid (TRPV) channels play a critical role in calcium homeostasis, pain sensation, immunological response, and cancer progression. TRPV channels are blocked by ruthenium red (RR), a universal pore blocker for a wide array of cation channels. Here we use cryo-electron microscopy to reveal the molecular details of RR block in TRPV2 and TRPV5, members of the two TRPV subfamilies. In TRPV2 activated by 2-aminoethoxydiphenyl borate, RR is tightly coordinated in the open selectivity filter, blocking ion flow and preventing channel inactivation. In TRPV5 activated by phosphatidylinositol 4,5-bisphosphate, RR blocks the selectivity filter and closes the lower gate through an interaction with polar residues in the pore vestibule. Together, our results provide a detailed understanding of TRPV subfamily pore block, the dynamic nature of the selectivity filter and allosteric communication between the selectivity filter and lower gate. The cryo-EM structures of TRPV2 and TRPV5, members of the two TRPV subfamilies, in complex with the ruthenium red blocker, reveal the intimate relationship between channel gating and pore blocking.The ruthenium red is a pore blocker in both subfamilies of the TRPV channels. In TRPV2 activated by 2-APB, ruthenium red is tightly coordinated in the open selectivity filter and prevents channel inactivation through allosteric mechanism. In TRPV5 activated by PI(4,5)P2, the ruthenium red in the selectivity filter closes the lower gate by interacting with polar residues in the pore vestibule. The cryo-EM structures of TRPV2 and TRPV5, members of the two TRPV subfamilies, in complex with the ruthenium red blocker, reveal the intimate relationship between channel gating and pore blocking.