Tweety-homolog (Ttyh) Family Encodes the Pore-forming Subunits of the Swelling-dependent Volume-regulated Anion Channel (VRACswell) in the Brain

Young-Eun Han; Jea Kwon; Joungha Won; Heeyoung An; Minwoo Wendy Jang; Junsung Woo; Je Sun Lee; Min Gu Park; Bo-Eun Yoon; Seung Eun Lee; Eun Mi Hwang; Jae-Young Jung; Hyungju Park; Soo-Jin Oh; C Justin Lee

doi:10.5607/en.2019.28.2.183

Tweety-homolog (Ttyh) Family Encodes the Pore-forming Subunits of the Swelling-dependent Volume-regulated Anion Channel (VRAC_swell) in the Brain

Exp Neurobiol. 2019 Apr;28(2):183-215. doi: 10.5607/en.2019.28.2.183. Epub 2019 Apr 30.

Authors

Young-Eun Han^{1

2

3}, Jea Kwon^{1

3

4}, Joungha Won^{1

3

5}, Heeyoung An^{1

3

4}, Minwoo Wendy Jang^{1

3

4}, Junsung Woo³, Je Sun Lee⁶, Min Gu Park^{1

3

4}, Bo-Eun Yoon⁷, Seung Eun Lee⁸, Eun Mi Hwang³, Jae-Young Jung^{2

3}, Hyungju Park⁶, Soo-Jin Oh^{3

9}, C Justin Lee^{1

2

3}

Affiliations

¹ Center for Cognition and Sociality, Institute for Basic Science, Daejeon 34126, Korea.
² Department of Neuroscience, Division of Bio-Medical Science & Technology, KIST School, Korea University of Science and Technology, Seoul 02792, Korea.
³ Center for Glia-Neuron Interaction, Korea Institute of Science and Technology (KIST), Seoul 02792, Korea.
⁴ KU-KIST, Graduate School of Converging Science and Technology, Korea University, Seoul, 02841, Korea.
⁵ Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 34141, Korea.
⁶ Molecular Neurobiology Laboratory, Dept. of Structure and Function of Neural Network, Korea Brain Research Institute, Daegu 41068, Korea.
⁷ Department of molecular biology, Dankook University, Cheonan 31116, Korea.
⁸ Virus Facility, Research Animal Resource Center, Korea Institute of Science and Technology (KIST), Seoul 02792, Korea.
⁹ Convergence Research Center for Diagnosis, Treatment and Care System of Dementia, Korea Institute of Science and Technology (KIST), Seoul 02792, Korea.

Abstract

In the brain, a reduction in extracellular osmolality causes water-influx and swelling, which subsequently triggers Cl^-- and osmolytes-efflux via volume-regulated anion channel (VRAC). Although LRRC8 family has been recently proposed as the pore-forming VRAC which is activated by low cytoplasmic ionic strength but not by swelling, the molecular identity of the pore-forming swelling-dependent VRAC (VRAC_swell) remains unclear. Here we identify and characterize Tweety-homologs (TTYH1, TTYH2, TTYH3) as the major VRAC_swell in astrocytes. Gene-silencing of all Ttyh1/2/3 eliminated hypo-osmotic-solution-induced Cl^- conductance (I_Cl,swell) in cultured and hippocampal astrocytes. When heterologously expressed in HEK293T or CHO-K1 cells, each TTYH isoform showed a significant I_Cl,swell with similar aquaporin-4 dependency, pharmacological properties and glutamate permeability as I_Cl,swell observed in native astrocytes. Mutagenesis-based structure-activity analysis revealed that positively charged arginine residue at 165 in TTYH1 and 164 in TTYH2 is critical for the formation of the channel-pore. Our results demonstrate that TTYH family confers the bona fide VRAC_swell in the brain.

Keywords: Ttyh; Tweety-homolog; VRAC; Volume regulation; Volume-regulated anion channel.