Cryo-EM structures of an LRRC8 chimera with native functional properties reveal heptameric assembly

Volume-regulated anion channels (VRACs) mediate volume regulatory Cl- and organic solute efflux from vertebrate cells. VRACs are heteromeric assemblies of LRRC8A-E proteins with unknown stoichiometries. Homomeric LRRC8A and LRRC8D channels have a small pore, hexameric structure. However, these channels are either non-functional or exhibit abnormal regulation and pharmacology, limiting their utility for structure-function analyses. We circumvented these limitations by developing novel homomeric LRRC8 chimeric channels with functional properties consistent with those of native VRAC/LRRC8 channels. We demonstrate here that the LRRC8C-LRRC8A(IL1(25)) chimera comprising LRRC8C and 25 amino acids unique to the first intracellular loop (IL1) of LRRC8A has a heptameric structure like that of homologous pannexin channels. Unlike homomeric LRRC8A and LRRC8D channels, heptameric LRRC8C-LRRC8A(IL1(25)) channels have a large-diameter pore similar to that estimated for native VRACs, exhibit normal DCPIB pharmacology, and have higher permeability to large organic anions. Lipid-like densities are located between LRRC8C-LRRC8A(IL1(25)) subunits and occlude the channel pore. Our findings provide new insights into VRAC/LRRC8 channel structure and suggest that lipids may play important roles in channel gating and regulation.

Automated summary

Volume-regulated anion channels (VRACs) mediate volume regulation and solute efflux from cells, but their composition and structure remain unclear. In this study, the authors developed novel homomeric LRRC8 chimeric channels that mimic native VRACs. They found that the LRRC8C-LRRC8A(IL1(25)) chimera has a seven-subunit structure, a large pore size, and normal pharmacology compared to non-functional homomeric channels. Lipid-like densities were also observed in the channel, suggesting a role in gating and regulation.