LRRC8 family proteins within lysosomes regulate cellular osmoregulation and enhance cell survival to multiple physiological stresses

Proc Natl Acad Sci U S A. 2020 Nov 17;117(46):29155-29165. doi: 10.1073/pnas.2016539117.

Ping Li ¹ ², Meiqin Hu ³ ², Ce Wang ³, Xinghua Feng ², ZhuangZhuang Zhao ², Ying Yang ³, Nirakar Sahoo ³ ⁴, Mingxue Gu ³, Yexin Yang ³, Shiyu Xiao ³, Rajan Sah ⁵, Timothy L Cover ⁶ ⁷ ⁸, Janet Chou ⁹, Raif Geha ⁹, Fernando Benavides ¹⁰, Richard I Hume ¹, Haoxing Xu ³

Affiliations

1 Department of Molecular, Cellular, and Developmental Biology, The University of Michigan, Ann Arbor, MI 48109-1085; pinglium@umich.edu rhume@umich.edu.
2 Collaborative Innovation Center of Yangtze River Delta Region Green Pharmaceuticals, College of Pharmaceutical Sciences, Zhejiang University of Technology, 310014 Hangzhou, China.
3 Department of Molecular, Cellular, and Developmental Biology, The University of Michigan, Ann Arbor, MI 48109-1085.
4 Department of Biology, The University of Texas Rio Grande Valley, Edinburg, TX 78539-2999.
5 Center for Cardiovascular Research, Division of Cardiology, Department of Internal Medicine, Washington University School of Medicine, St. Louis, MO 63110.
6 Department of Medicine, Vanderbilt University School of Medicine, Nashville, TN 37212.
7 Department of Pathology, Microbiology and Immunology, Vanderbilt University School of Medicine, Nashville, TN 37212.
8 Veterans Affairs Tennessee Valley Healthcare System, Nashville, TN 37212.
9 Division of Immunology, Boston Children's Hospital, Boston, MA 02115.
10 Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, TX 78957.

PMID: 33139539 DOI: 10.1073/pnas.2016539117

Abstract

LRRC8 family proteins on the plasma membrane play a critical role in cellular osmoregulation by forming volume-regulated anion channels (VRACs) necessary to prevent necrotic cell death. We demonstrate that intracellular LRRC8 proteins acting within lysosomes also play an essential role in cellular osmoregulation. LRRC8 proteins on lysosome membranes generate large lysosomal volume-regulated anion channel (Lyso-VRAC) currents in response to low cytoplasmic ionic strength conditions. When a double-leucine L⁷⁰⁶L⁷⁰⁷ motif at the C terminus of LRRC8A was mutated to alanines, normal plasma membrane VRAC currents were still observed, but Lyso-VRAC currents were absent. We used this targeting mutant, as well as pharmacological tools, to demonstrate that Lyso-VRAC currents are necessary for the formation of large lysosome-derived vacuoles, which store and then expel excess water to maintain cytosolic water homeostasis. Thus, Lyso-VRACs allow lysosomes of mammalian cells to act as the cell`s "bladder." When Lyso-VRAC current was selectively eliminated, the extent of necrotic cell death to sustained stress was greatly increased, not only in response to hypoosmotic stress, but also to hypoxic and hypothermic stresses. Thus Lyso-VRACs play an essential role in enabling cells to mount successful homeostatic responses to multiple stressors.

Keywords

chloride channel; exocytosis; lysosome; osmoregulation; vacuolation.

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