Cellular volume regulation by anoctamin 6: Ca2+, phospholipase A2 and osmosensing
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During cell swelling, Cl− channels are activated to lower intracellular Cl− concentrations and to reduce cell volume, a process termed regulatory volume decrease (RVD). We show that anoctamin 6 (ANO6; TMEM16F) produces volume-regulated anion currents and controls cell volume in four unrelated cell types. Volume regulation is compromised in freshly isolated intestinal epithelial cells from Ano6−/− mice and also in lymphocytes from a patient lacking expression of ANO6. Ca2+ influx is activated and thus ANO6 is stimulated during cell swelling by local Ca2+ increase probably in functional nanodomains near the plasma membrane. This leads to stimulation of phospholipase A2 (PLA2) and generation of plasma membrane lysophospholipids, which activates ANO6. Direct application of lysophospholipids also activates an anion current that is inhibited by typical ANO6 blocker. An increase in intracellular Ca2+ supports activation of ANO6, but is not required when PLA2 is fully activated, while re-addition of arachidonic acid completely blocked ANO6. Moreover, ANO6 is activated by low intracellular Cl− concentrations and may therefore operate as a cellular osmosensor. High intracellular Cl− concentration inhibits ANO6 and activation by PLA2. Taken together, ANO6 supports volume regulation and volume activation of anion currents by action as a Cl− channel or by scrambling membrane phospholipids. Thereby, it may support the function of LRRC8 proteins.
KeywordsTMEM16F Anoctamin 6 Volume regulation Regulatory volume decrease RVD Volume-regulated anion channel VRAC Apoptosis
This study was supported by DFG SFB699-A7/A12, DFG KU756/12-1, Sander-Stiftung 2013.031.1 and Volkswagenstiftung AZ 87 499. We thank Dr. Johan Heemskerk and Dr. Eduard Bevers for supplying the lymphocyte cell lines. The excellent technical assistance by Mss. B. Wild, P. Seeberger, E. Tartler, Inês Cabrita and Mr. Simon Höllerer is gratefully acknowledged.
Author contribution statement
L.S., J.O., P.W. and R.S. performed the experiments. L.S., J.O., P.W., R.S. and K.K. wrote the manuscript and prepared the figures. L.S., J.O., P.W., R.S. and K.K. reviewed the final manuscript.
Compliance with ethical standards
All animal experiments were approved by the local ethics commission of the University of Regensburg and were conducted according to the guidelines of the American Physiological Society and the German law for welfare of animals.
Conflict of interest
The authors declare that they have no competing interests.
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