Functional Characterization of Leucine‐Rich Repeat Containing 8 A (LRRC8A) Homomeric Channel

Abstract

Regulation of cell volume in response to osmotic perturbations is an essential cellular process. The volume‐regulated anion channel (VRAC) is ubiquitously expressed in vertebrate cells and mediates the swelling‐induced release of chloride and organic solutes. VRAC is encoded by LRRC8 gene family members, LRRC8A‐E. VRAC is a heteromeric channel comprising an essential LRRC8A subunit plus at least one other LRRC8 member. Several high‐resolution cryo‐EM structures of the LRRC8A homomeric channel have recently been published. However, until now, the detailed functional and regulatory properties of the LRRC8A homomeric channel have not yet been characterized. Here, we show that HCT116 cells in which all five Lrrc8 genes have been knocked out produce a small whole‐cell current (~20 pA/pF at +100 mV) when transfected with a plasmid encoding human LRRC8A. In striking contrast to the heteromeric VRAC current, the LRRC8A current is not activated by cell swelling or inhibited by cell shrinkage. However, when cells are patch clamped with an intracellular solution of low ionic strength, the current exhibits slow and modest activation, and is dramatically activated by cell swelling. The degree of swelling‐induced LRRC8A current activation is directly and dose‐dependently related to intracellular ionic strength. Consistent with the heteromeric VRAC current, swelling‐induced activation of LRRC8A current is prevented by removing intracellular ATP. We are currently evaluating the anion selectivity and pharmacological properties of LRRC8A. Our studies provide important new insights into the functional and regulatory properties of the LRRC8A homomeric channels.Support or Funding InformationThis work was funded by National Institutes of Health grant R01DK051610 to K.S.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.