E. K. Hoffmann, I. H. Lambert, and P. Sf, Physiology of cell volume regulation in vertebrates, Physiol Rev, vol.89, issue.1, pp.193-277, 2009.

R. E. Day, P. Kitchen, D. S. Owen, C. Bland, L. Marshall et al., Human aquaporins: regulators of transcellular water flow, Biochim Biophys Acta, vol.1840, issue.5, pp.1492-506, 2014.

C. H. Yeung, J. P. Barfield, and C. Tg, Chloride channels in physiological volume regulation of human spermatozoa, Biol Reprod, vol.73, issue.5, pp.1057-63, 2005.

T. G. Cooper and C. H. Yeung, Involvement of potassium and chloride channels and other transporters in volume regulation by spermatozoa, Curr Pharm Des, vol.13, issue.31, pp.3222-3252, 2007.

T. J. Jentsch, D. Lutter, R. Planells-cases, F. Ullrich, and F. K. Voss, VRAC: molecular identification as LRRC8 heteromers with differential functions, Pflugers Arch, vol.468, issue.3, pp.385-93, 2016.

S. F. Pedersen, T. K. Klausen, and B. Nilius, The identification of a volume-regulated anion channel: an amazing Odyssey, Acta Physiol (Oxf), vol.213, issue.4, pp.868-81, 2015.

R. Syeda, Z. Qiu, A. E. Dubin, S. E. Murthy, M. N. Florendo et al.,

, Proteins Form Volume-Regulated Anion Channels that Sense Ionic Strength

, Cell, vol.164, issue.3, pp.499-511, 2016.

D. D. Mruk and C. Y. Cheng, Tight junctions in the testis: new perspectives, Philos Trans R Soc Lond B Biol Sci, vol.365, pp.1621-1656, 1546.

T. G. Cooper, C. H. Yeung, A. Wagenfeld, E. Nieschlag, M. Poutanen et al., Mouse models of infertility due to swollen spermatozoa, Mol Cell Endocrinol, vol.216, issue.1-2, pp.55-63, 2004.

A. M. Petrunkina, R. A. Harrison, M. Ekhlasi-hundrieser, and E. Topfer-petersen, Role of volume-stimulated osmolyte and anion channels in volume regulation by mammalian sperm, Mol Hum Reprod, vol.10, issue.11, pp.815-838, 2004.

C. H. Yeung, J. P. Barfield, and C. Tg, Physiological volume regulation by spermatozoa, Mol Cell Endocrinol, vol.250, issue.1-2, pp.98-105, 2006.

C. H. Yeung, M. Anapolski, P. Sipila, A. Wagenfeld, M. Poutanen et al., Sperm volume regulation: maturational changes in fertile and infertile transgenic mice and association with kinematics and tail angulation, Biol Reprod, vol.67, issue.1, pp.269-75, 2002.

R. F. Domeniconi, A. C. Souza, B. Xu, A. M. Washington, and H. Bt, Is the Epididymis a Series of Organs Placed Side By Side?, Biol Reprod, vol.95, issue.1, p.10, 2016.

C. H. Yeung, C. Callies, A. Rojek, S. Nielsen, and C. Tg, Aquaporin isoforms involved in physiological volume regulation of murine spermatozoa, Biol Reprod, vol.80, issue.2, pp.350-357, 2009.

G. Smits and A. V. Kajava, LRRC8 extracellular domain is composed of 17 leucinerich repeats, Mol Immunol, vol.41, issue.5, pp.561-563, 2004.

F. Abascal and R. Zardoya, LRRC8 proteins share a common ancestor with pannexins, and may form hexameric channels involved in cell-cell communication, Bioessays, vol.34, issue.7, pp.551-60, 2012.

Y. Zhang, L. Xie, S. K. Gunasekar, D. Tong, A. Mishra et al., SWELL1 is a regulator of adipocyte size, insulin signalling and glucose homeostasis, Nat Cell Biol, vol.19, issue.5, pp.504-521, 2017.

C. C. Lee, E. Freinkman, D. M. Sabatini, and P. Hl, The protein synthesis inhibitor blasticidin s enters mammalian cells via leucine-rich repeat-containing protein 8D, J Biol Chem, vol.289, issue.24, pp.17124-17155, 2014.

D. Lutter, F. Ullrich, J. C. Lueck, S. Kempa, and J. Tj, Selective transport of neurotransmitters and modulators by distinct volume-regulated LRRC8 anion channels, J Cell Sci, vol.130, issue.6, pp.1122-1155, 2017.

C. Kang, L. Xie, S. K. Gunasekar, A. Mishra, Y. Zhang et al., SWELL1 is a glucose sensor regulating beta-cell excitability and systemic glycaemia, Nat Commun, vol.9, issue.1, p.367, 2018.

A. Lalouette, A. Lablack, J. L. Guenet, X. Montagutelli, and D. Segretain, Male sterility caused by sperm cell-specific structural abnormalities in ebouriffe, a new mutation of the house mouse, Biol Reprod, vol.55, issue.2, pp.355-63, 1996.

C. D. Platt, J. Chou, P. Houlihan, Y. R. Badran, L. Kumar et al., Leucinerich repeat containing 8A (LRRC8A)-dependent volume-regulated anion channel activity is dispensable for T-cell development and function, J Allergy Clin Immunol, vol.140, issue.6, pp.1651-1660, 2017.

L. Kumar, J. Chou, C. S. Yee, A. Borzutzky, E. H. Vollmann et al.,

, Consortium GT. Human genomics. The Genotype-Tissue Expression (GTEx) pilot analysis: multitissue gene regulation in humans, J Exp Med, vol.211, issue.5, pp.648-60, 2014.

G. Margolin, P. P. Khil, J. Kim, M. A. Bellani, and C. Rd, Integrated transcriptome analysis of mouse spermatogenesis, BMC Genomics, vol.15, p.39, 2014.

W. Li, J. Wu, S. Y. Kim, M. Zhao, S. A. Hearn et al., Chd5 orchestrates chromatin remodelling during sperm development, Nat Commun, vol.5, p.3812, 2014.

J. Bao, S. Rousseaux, J. Shen, K. Lin, Y. Lu et al., The arginine methyltransferase CARM1 represses p300*ACT*CREMtau activity and is required for spermiogenesis, Nucleic Acids Res, 2018.

N. Kotaja, S. Kimmins, S. Brancorsini, D. Hentsch, J. L. Vonesch et al.,

, Preparation, isolation and characterization of stage-specific spermatogenic cells for cellular and molecular analysis, Nat Methods, vol.1, issue.3, pp.249-54, 2004.

J. Bao, K. Vitting-seerup, J. Waage, C. Tang, Y. Ge et al., UPF2-Dependent Nonsense-Mediated mRNA Decay Pathway Is Essential for Spermatogenesis by Selectively Eliminating Longer 3'UTR Transcripts, PLoS Genet, vol.12, issue.5, p.1005863, 2016.

C. Coutton, J. Escoffier, G. Martinez, C. Arnoult, and R. Pf, Teratozoospermia: spotlight on the main genetic actors in the human, Hum Reprod Update, vol.21, issue.4, pp.455-85, 2015.

C. Krausz and R. , Genetics of male infertility, Nat Rev Urol, 2018.

C. Dong, P. Wei, X. Jian, R. Gibbs, E. Boerwinkle et al., Comparison and integration of deleteriousness prediction methods for nonsynonymous SNVs in whole exome sequencing studies, Hum Mol Genet, vol.24, issue.8, pp.2125-2162, 2015.

M. Biasini, S. Bienert, A. Waterhouse, K. Arnold, G. Studer et al.,

, SWISS-MODEL: modelling protein tertiary and quaternary structure using evolutionary information, Nucleic Acids Res, vol.42, pp.252-260, 2014.

H. Gaitan-penas, A. Gradogna, L. Laparra-cuervo, C. Solsona, V. Fernandez-duenas et al., Investigation of LRRC8-Mediated Volume-Regulated Anion Currents in Xenopus Oocytes, Biophys J, vol.111, issue.7, pp.1429-1472, 2016.

H. Gaitan-penas, M. Pusch, and R. Estevez, Expression of LRRC8/VRAC Currents in Xenopus Oocytes: Advantages and Caveats, Int J Mol Sci, vol.19, issue.3, 2018.

C. H. Yeung, J. P. Barfield, and C. Tg, The role of anion channels and Ca2+ in addition to K+ channels in the physiological volume regulation of murine spermatozoa, Mol Reprod Dev, vol.71, issue.3, pp.368-79, 2005.

H. Qi, M. M. Moran, B. Navarro, J. A. Chong, G. Krapivinsky et al., All four CatSper ion channel proteins are required for male fertility and sperm cell hyperactivated motility, Proc Natl Acad Sci, vol.104, issue.4, pp.1219-1242, 2007.

X. H. Zeng, C. Yang, S. T. Kim, C. J. Lingle, and X. M. Xia, Deletion of the Slo3 gene abolishes alkalization-activated K+ current in mouse spermatozoa, Proc Natl Acad Sci, vol.108, issue.14, pp.5879-84, 2011.

, LacZ/Neo cassette abolished Lrrc8a gene expression. (B) Sperm harboring the targeted conditional Lrrc8a flox allele

K. O. Lrrc8a, cKO) mice, by breeding with a germline-specific cre line

, Representative morphology of sperm visualized by phase contrast (top, bar=3 µm) and SEM (bottom, bar=2 µm), retrieved from the cauda epididymis in the WT, Lrrc8a F443*/F443* , KO and cKO mice. Figure 6. A hypomorphic mutation (c.1634G>A, R545H) in the LRRC8A gene identified in a male sterile patient, Lrrc8a fl/ ? , cKO). (C)

, LRRC8A R545H ) (bottom) in a male patient suffering from infertility. (B) Sanger re-sequencing validated the c.1634G>A mutation in this patient. A red rectangle highlights the mutation site. (C) HE staining of testis biopsies from the patient (#90) and a fertile control. Spg, Spermatogonia. Spc, Spermatocyte. Spd, Spermatid. Bar=15 µm. (D) 3-Dimensional homology modeling of protein structure for WT (top) and mutant LRRC8A R545H protein (bottom), Whole Exome Sequencing (WES) identified a missense point mutation (c.1634G>A, p.Arg545His) in a highly conserved region of LRRC8A (top), resulting in the substitution of arginine (R) to histidine (H)

, Three batches of cRNAs were injected into 15 oocytes for each group in each assay. Representative western blot performed using the same injected oocytes to demonstrate comparable expression levels of the injected cRNAs (F, right panel). Black dots indicate the expected size of fluorescently tagged WT or mutant LRRC8A. Comparison of the normalized slope conductance induced by coexpressing 8A HA-VFP and 8A(R545H) HA-VFP with 8C-mCherry or 8D-mCherry as indicated (G, left panel). ELISA-based luminescence assays utilizing the HA tag illustrated comparable expression of the injected, temdomly tagged cRNAs in the oocyte membranes (G, right panel). Three batches of cRNAs were injected into 15 oocytes for each group in each assay. PM, Plasma membrane expression. Data represent mean ± SD of three independent experiments with 3 batches of cRNA preparations, traces of oocytes are shown for the co-injections of WT LRRC8A-VFP (8A-VFP) or LRRC8A R545H -VFP [8A(R545H)-VFP] with LRRC8C-mCherry (8C-mCh) or LRRC8D-mCherry (8D-mCh) as labeled (E) (n=30)