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Electrophysiological Techniques on the Study of Endolysosomal Ion Channels

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Endolysosomal Voltage-Dependent Cation Channels

Part of the book series: Handbook of Experimental Pharmacology ((HEP,volume 278))

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Abstract

Endolysosomal ion channels are a group of ion channel proteins that are functionally expressed on the membrane of endolysosomal vesicles. The electrophysiological properties of these ion channels in the intracellular organelle membrane cannot be observed using conventional electrophysiological techniques. This section compiles the different electrophysiological techniques utilized in recent years to study endolysosomal ion channels and describes their methodological characteristics, emphasizing the most widely used technique for whole endolysosome recordings to date. This includes the use of different pharmacological tools and genetic tools for the application of patch-clamping techniques for specific stages of endolysosomes, allowing the recording of ion channel activity in different organelles, such as recycling endosomes, early endosomes, late endosomes, and lysosomes. These electrophysiological techniques are not only cutting-edge technologies that help to investigate the biophysical properties of known and unknown intracellular ion channels but also help us to investigate the physiopathological role of these ion channels in the distribution of dynamic vesicles and to identify new therapeutic targets for precision medicine and drug screening.

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References

  • Bertl A, Blumwald E, Coronado R, Eisenberg R, Findlay G, Gradmann D, Hille B, Köhler K, Kolb HA, MacRobbie E et al (1992) Electrical measurements on endomembranes. Science 258(5084):873–874

    Article  CAS  PubMed  Google Scholar 

  • Bobak N, Feliciangeli S, Chen CC, Ben Soussia I, Bittner S, Pagnotta S, Ruck T, Biel M, Wahl-Schott C, Grimm C, Meuth SG, Lesage F (2017) Recombinant tandem of pore-domains in a weakly inward rectifying K+ channel 2 (TWIK2) forms active lysosomal channels. Sci Rep 7(1):649

    Article  PubMed  PubMed Central  Google Scholar 

  • Bonifacino JS, Traub LM (2003) Signals for sorting of transmembrane proteins to endosomes and lysosomes. Annu Rev Biochem 72:395–447

    Article  CAS  PubMed  Google Scholar 

  • Brailoiu E, Hooper R, Cai X, Brailoiu GC, Keebler MV, Dun NJ, Marchant JS, Patel S (2010) An ancestral deuterostome family of two-pore channels mediates nicotinic acid adenine dinucleotide phosphate-dependent calcium release from acidic organelles. J Biol Chem 285(5):2897–2901

    Article  CAS  PubMed  Google Scholar 

  • Cai X, Xu Y, Kim YM, Loureiro J, Huang Q (2014) PIKfyve, a class III lipid kinase, is required for TLR-induced type I IFN production via modulation of ATF3. J Immunol 192(7):3383–3389

    Article  CAS  PubMed  Google Scholar 

  • Cang C, Zhou Y, Navarro B, Seo YJ, Aranda K, Shi L, Battaglia-Hsu S, Nissim I, Clapham DE, Ren D (2013) mTOR regulates lysosomal ATP-sensitive two-pore Na(+) channels to adapt to metabolic state. Cell 152(4):778–790

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Cang C, Aranda K, Seo YJ, Gasnier B, Ren D (2015) TMEM175 is an organelle K(+) channel regulating lysosomal function. Cell 162(5):1101–1112

    Article  CAS  PubMed  Google Scholar 

  • Cao Q, Zhong XZ, Zou Y, Zhang Z, Toro L, Dong XP (2015) BK channels alleviate lysosomal storage diseases by providing positive feedback regulation of lysosomal Ca2+ release. Dev Cell 33(4):427–441

    Article  CAS  PubMed  Google Scholar 

  • Carpentier S, N'Kuli F, Grieco G, Van Der Smissen P, Janssens V, Emonard H, Bilanges B, Vanhaesebroeck B, Gaide Chevronnay HP, Pierreux CE, Tyteca D, Courtoy PJ (2013) Class III phosphoinositide 3-kinase/VPS34 and dynamin are critical for apical endocytic recycling. Traffic 14(8):933–948

    Article  CAS  PubMed  Google Scholar 

  • Cerny J, Feng Y, Yu A, Miyake K, Borgonovo B, Klumperman J, Meldolesi J, McNeil PL, Kirchhausen T (2004) The small chemical vacuolin-1 inhibits Ca(2+)-dependent lysosomal exocytosis but not cell resealing. EMBO Rep 5(9):883–888

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chao YK, Schludi V, Chen CC, Butz E, Nguyen ONP, Müller M, Krüger J, Kammerbauer C, Ben-Johny M, Vollmar AM, Berking C, Biel M, Wahl-Schott CA, Grimm C (2017) TPC2 polymorphisms associated with a hair pigmentation phenotype in humans result in gain of channel function by independent mechanisms. Proc Natl Acad Sci U S A 114(41):E8595–E8602

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chen CC, Keller M, Hess M, Schiffmann R, Urban N, Wolfgardt A, Schaefer M, Bracher F, Biel M, Wahl-Schott C, Grimm C (2014) A small molecule restores function to TRPML1 mutant isoforms responsible for mucolipidosis type IV. Nat Commun 5:4681

    Article  CAS  PubMed  Google Scholar 

  • Chen CC, Cang C, Fenske S, Butz E, Chao YK, Biel M, Ren D, Wahl-Schott C, Grimm C (2017a) Patch-clamp technique to characterize ion channels in enlarged individual endolysosomes. Nat Protoc 12(8):1639–1658

    Article  CAS  PubMed  Google Scholar 

  • Chen CC, Butz ES, Chao YK, Grishchuk Y, Becker L, Heller S, Slaugenhaupt SA, Biel M, Wahl-Schott C, Grimm C (2017b) Small molecules for early endosome-specific patch clamping. Cell Chem Biol 24(7):907–916.e4

    Article  CAS  PubMed  Google Scholar 

  • Chen Q, Kang J, Fu C (2018) The independence of and associations among apoptosis, autophagy, and necrosis. Signal Transduct Target Ther 3:18

    Article  PubMed  PubMed Central  Google Scholar 

  • Chen CC, Krogsaeter E, Butz ES, Li Y, Puertollano R, Wahl-Schott C, Biel M, Grimm C (2020) TRPML2 is an osmo/mechanosensitive cation channel in endolysosomal organelles. Sci Adv 6(46):eabb5064

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chen CC, Krogsaeter E, Kuo CY, Huang MC, Chang SY, Biel M (2022) Endolysosomal cation channels point the way towards precision medicine of cancer and infectious diseases. Biomed Pharmacother 148:112751

    Article  CAS  PubMed  Google Scholar 

  • de Duve C, Pressman BC, Gianetto R, Wattiaux R, Appelmans F (1955) Tissue fractionation studies. 6. Intracellular distribution patterns of enzymes in rat-liver tissue. Biochem J 60:604–617

    Article  PubMed  PubMed Central  Google Scholar 

  • Dong XP, Shen D, Wang X, Dawson T, Li X, Zhang Q, Cheng X, Zhang Y, Weisman LS, Delling M, Xu H (2010) PI(3,5)P(2) controls membrane trafficking by direct activation of mucolipin Ca(2+) release channels in the endolysosome. Nat Commun 1(4):38

    Article  PubMed  Google Scholar 

  • Escobar L, Echeverria OM, Vazquez-Nin GH (2015) Necrosis as programmed cell death. https://www.intechopen.com/books/cell-death-autophagy-apoptosis-and-necrosis/necrosis-as-programmed-cell-death

  • Freeman SA, Grinstein S (2018) Resolution of macropinosomes, phagosomes and autolysosomes: osmotically driven shrinkage enables tubulation and vesiculation. Traffic 19(12):965–974

    Article  CAS  PubMed  Google Scholar 

  • Gerndt S, Chen CC, Chao YK, Yuan Y, Burgstaller S, Scotto Rosato A, Krogsaeter E, Urban N, Jacob K, Nguyen ONP, Miller MT, Keller M, Vollmar AM, Gudermann T, Zierler S, Schredelseker J, Schaefer M, Biel M, Malli R, Wahl-Schott C, Bracher F, Patel S, Grimm C (2020) Agonist-mediated switching of ion selectivity in TPC2 differentially promotes lysosomal function. Elife 9:e54712

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Greene W, Gao SJ (2009) Actin dynamics regulate multiple endosomal steps during Kaposi’s sarcoma-associated herpesvirus entry and trafficking in endothelial cells. PLoS Pathog 5(7):e1000512

    Article  PubMed  PubMed Central  Google Scholar 

  • Grimm C, Holdt LM, Chen CC, Hassan S, Müller C, Jörs S, Cuny H, Kissing S, Schröder B, Butz E, Northoff B, Castonguay J, Luber CA, Moser M, Spahn S, Lüllmann-Rauch R, Fendel C, Klugbauer N, Griesbeck O, Haas A, Mann M, Bracher F, Teupser D, Saftig P, Biel M, Wahl-Schott C (2014) High susceptibility to fatty liver disease in two-pore channel 2-deficient mice. Nat Commun 5:4699

    Article  CAS  PubMed  Google Scholar 

  • Hata Y, Slaughter CA, Südhof TC (1993) Synaptic vesicle fusion complex containsunc-18 homologue bound to syntaxin. Nature 366(6453):347–351

    Article  CAS  PubMed  Google Scholar 

  • Hu M, Zhou N, Cai W, Xu H (2022) Lysosomal solute and water transport. J Cell Biol 221(11):e202109133

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Huang P, Zou Y, Zhong XZ, Cao Q, Zhao K, Zhu MX, Murrell-Lagnado R, Dong XP (2014) P2X4 forms functional ATP-activated cation channels on lysosomal membranes regulated by luminal pH. J Biol Chem 289(25):17658–17667

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Huotari J, Helenius A (2011) Endosome maturation. EMBO J 30(17):3481–3500

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Izu YC, Sachs F (1991) Inhibiting synthesis of extracellular matrix improves patch clamp seal formation. Pflugers Arch 419(2):218–220

    Article  CAS  PubMed  Google Scholar 

  • Kaiser CA, Schekman R (1990) Distinct sets of SEC genes govern transport vesicle formation and fusion early in the secretory pathway. Cell 61(4):723–733

    Article  CAS  PubMed  Google Scholar 

  • Karvar S, Yao X, Crothers JM, Liu Y, Forte JG (2002) Localization and function of soluble N-ethylmaleimide-sensitive factor attachment protein-25 and vesicle-associated membrane protein-2 in functioning gastric parietal cells. J Biol Chem 277(51):50030–50035

    Article  CAS  PubMed  Google Scholar 

  • Klionsky DJ, Eskelinen EL, Deretic V (2014) Autophagosomes, phagosomes, autolysosomes, phagolysosomes, autophagolysosomes... wait, I’m confused. Autophagy 10(4):549–551

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Klumperman J, Raposo G (2014) The complex ultrastructure of the endolysosomal system. Cold Spring Harb Perspect Biol 6(10):a016857

    Article  PubMed  PubMed Central  Google Scholar 

  • Li X, Wang X, Zhang X, Zhao M, Tsang WL, Zhang Y, Yau RG, Weisman LS, Xu H (2013) Genetically encoded fluorescent probe to visualize intracellular phosphatidylinositol 3,5-bisphosphate localization and dynamics. Proc Natl Acad Sci U S A 110(52):21165–21170

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Li P, Hu M, Wang C, Feng X, Zhao Z, Yang Y, Sahoo N, Gu M, Yang Y, Xiao S, Sah R, Cover TL, Chou J, Geha R, Benavides F, Hume RI, Xu H (2020) LRRC8 family proteins within lysosomes regulate cellular osmoregulation and enhance cell survival to multiple physiological stresses. Proc Natl Acad Sci U S A 117(46):29155–29165

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Li Y, Schön C, Chen CC, Yang Z, Liegl R, Murenu E, Schworm B, Klugbauer N, Grimm C, Wahl-Schott C, Michalakis S, Biel M (2021) TPC2 promotes choroidal angiogenesis and inflammation in a mouse model of neovascular age-related macular degeneration. Life Sci Alliance 4(8):e202101047

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lin-Moshier Y, Keebler MV, Hooper R, Boulware MJ, Liu X, Churamani D, Abood ME, Walseth TF, Brailoiu E, Patel S, Marchant JS (2014) The two-pore channel (TPC) interactome unmasks isoform-specific roles for TPCs in endolysosomal morphology and cell pigmentation. Proc Natl Acad Sci U S A 111(36):13087–13092

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Luzio JP, Pryor PR, Bright NA (2007 Aug) Lysosomes: fusion and function. Nat Rev Mol Cell Biol 8(8):622–632

    Article  CAS  PubMed  Google Scholar 

  • Maxfield FR, McGraw TE (2004) Endocytic recycling. Nat Rev Mol Cell Biol 5(2):121–132

    Article  CAS  PubMed  Google Scholar 

  • Morgan AJ, Platt FM, Lloyd-Evans E, Galione A (2011) Molecular mechanisms of endolysosomal Ca2+ signalling in health and disease. Biochem J 439(3):349–374

    Article  CAS  PubMed  Google Scholar 

  • Naslavsky N, Caplan S (2018) The enigmatic endosome – sorting the ins and outs of endocytic trafficking. J Cell Sci 131(13):jcs216499

    Article  PubMed  PubMed Central  Google Scholar 

  • Novick P, Schekman R (1979) Secretion and cell-surface growth are blocked in a temperature-sensitive mutant of Saccharomyces cerevisiae. Proc Natl Acad Sci U S A 76(4):1858–1862

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Osei-Owusu J, Yang J, Leung KH, Ruan Z, Lü W, Krishnan Y, Qiu Z (2021) Proton-activated chloride channel PAC regulates endosomal acidification and transferrin receptor-mediated endocytosis. Cell Rep 34(4):108683

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Patel S, Yuan Y, Chen CC, Jaślan D, Gunaratne G, Grimm C, Rahman T, Marchant JS (2022) Electrophysiology of Endolysosomal two-pore channels: a current account. Cell 11(15):2368

    Article  CAS  Google Scholar 

  • Piper RC, Luzio P (2008) Late endosomes: sorting and partitioning in multivesicular bodies. Traffic 2(9):612–621

    Article  Google Scholar 

  • Pitt SJ, Funnell TM, Sitsapesan M, Venturi E, Rietdorf K, Ruas M, Ganesan A, Gosain R, Churchill GC, Zhu MX, Parrington J, Galione A, Sitsapesan R (2010) TPC2 is a novel NAADP-sensitive Ca2+ release channel, operating as a dual sensor of luminal pH and Ca2+. J Biol Chem 285(45):35039–35046

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Plesch E, Chen CC, Butz E, Scotto Rosato A, Krogsaeter EK, Yinan H, Bartel K, Keller M, Robaa D, Teupser D, Holdt LM, Vollmar AM, Sippl W, Puertollano R, Medina D, Biel M, Wahl-Schott C, Bracher F, Grimm C (2018) Selective agonist of TRPML2 reveals direct role in chemokine release from innate immune cells. Elife 7:e39720

    Article  PubMed  PubMed Central  Google Scholar 

  • Repnik U, Česen MH, Turk B (2013) The endolysosomal system in cell death and survival. Cold Spring Harb Perspect Biol 5(1):a008755

    Article  PubMed  PubMed Central  Google Scholar 

  • Riederer E, Cang C, Ren D (2022) Lysosomal ion channels: what are they good for and are they druggable targets? Annu Rev Pharmacol Toxicol

    Google Scholar 

  • Ruas M, Davis LC, Chen CC, Morgan AJ, Chuang KT, Walseth TF, Grimm C, Garnham C, Powell T, Platt N, Platt FM, Biel M, Wahl-Schott C, Parrington J, Galione A (2015) Expression of Ca2+-permeable two-pore channels rescues NAADP signalling in TPC-deficient cells. EMBO J 34(13):1743–1758

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ruivo R, Anne C, Sagné C, Gasnier B (2009) Molecular and cellular basis of lysosomal transmembrane protein dysfunction. Biochim Biophys Acta 1793(4):636–649

    Article  CAS  PubMed  Google Scholar 

  • Saftig P, Klumperman J (2009) Lysosome biogenesis and lysosomal membrane proteins: trafficking meets function. Nat Rev Mol Cell Biol 10(9):623–635

    Article  CAS  PubMed  Google Scholar 

  • Sahoo N, Gu M, Zhang X, Raval N, Yang J, Bekier M, Calvo R, Patnaik S, Wang W, King G, Samie M, Gao Q, Sahoo S, Sundaresan S, Keeley TM, Wang Y, Marugan J, Ferrer M, Samuelson LC, Merchant JL, Xu H (2017) Gastric acid secretion from parietal cells is mediated by a Ca2+ efflux channel in the tubulovesicle. Dev Cell 41(3):262–273.e6

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sakmann B, Neher E (1984) Patch clamp techniques for studying ionic channels in excitable membranes. Annu Rev Physiol 46:455–472

    Article  CAS  PubMed  Google Scholar 

  • Sakurai Y, Kolokoltsov AA, Chen CC, Tidwell MW, Bauta WE, Klugbauer N, Grimm C, Wahl-Schott C, Biel M, Davey RA (2015) Ebola virus. Two-pore channels control Ebola virus host cell entry and are drug targets for disease treatment. Science 347(6225):995–998

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Samie M, Wang X, Zhang X, Goschka A, Li X, Cheng X, Gregg E, Azar M, Zhuo Y, Garrity AG, Gao Q, Slaugenhaupt S, Pickel J, Zolov SN, Weisman LS, Lenk GM, Titus S, Bryant-Genevier M, Southall N, Juan M, Ferrer M, Xu H (2013) A TRP channel in the lysosome regulates large particle phagocytosis via focal exocytosis. Dev Cell 26(5):511–524

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schieder M, Rötzer K, Brüggemann A, Biel M, Wahl-Schott C (2010) Planar patch clamp approach to characterize ionic currents from intact lysosomes. Sci Signal 3(151):pl3

    Article  CAS  PubMed  Google Scholar 

  • Söllner T, Whiteheart SW, Brunner M, Erdjument-Bromage H, Geromanos S, Tempst P, Rothman JE (1993) SNAP receptors implicated in vesicle targeting and fusion. Nature 362(6418):318–324

    Article  PubMed  Google Scholar 

  • Sönnichsen B, De Renzis S, Nielsen E, Rietdorf J, Zerial M (2000) Distinct membrane domains on endosomes in the recycling pathway visualized by multicolor imaging of Rab4, Rab5, and Rab11. J Cell Biol 149(4):901–914

    Article  PubMed  PubMed Central  Google Scholar 

  • Spix B, Butz ES, Chen CC, Rosato AS, Tang R, Jeridi A, Kudrina V, Plesch E, Wartenberg P, Arlt E, Briukhovetska D, Ansari M, Günsel GG, Conlon TM, Wyatt A, Wetzel S, Teupser D, Holdt LM, Ectors F, Boekhoff I, Boehm U, García-Añoveros J, Saftig P, Giera M, Kobold S, Schiller HB, Zierler S, Gudermann T, Wahl-Schott C, Bracher F, Yildirim AÖ, Biel M, Grimm C (2022) Lung emphysema and impaired macrophage elastase clearance in mucolipin 3 deficient mice. Nat Commun 13(1):318

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Stenmark H (2009) Rab GTPases as coordinators of vesicle traffic. Nat Rev Mol Cell Biol 10(8):513–525

    Article  CAS  PubMed  Google Scholar 

  • Takeshige K, Baba M, Tsuboi S, Noda T, Ohsumi Y (1992) Autophagy in yeast demonstrated with proteinase-deficient mutants and conditions for its induction. J Cell Biol 119(2):301–311

    Article  CAS  PubMed  Google Scholar 

  • Wandinger-Ness A, Zerial M (2014) Rab proteins and the compartmentalization of the endosomal system. Cold Spring Harb Perspect Biol 6(11):a022616

    Article  PubMed  PubMed Central  Google Scholar 

  • Wang X, Zhang X, Dong XP, Samie M, Li X, Cheng X, Goschka A, Shen D, Zhou Y, Harlow J, Zhu MX, Clapham DE, Ren D, Xu H (2012) TPC proteins are phosphoinositide-activated sodium-selective ion channels in endosomes and lysosomes. Cell 151(2):372–383

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wang W, Zhang X, Gao Q, Lawas M, Yu L, Cheng X, Gu M, Sahoo N, Li X, Li P, Ireland S, Meredith A, Xu H (2017) A voltage-dependent K+ channel in the lysosome is required for refilling lysosomal Ca2+ stores. J Cell Biol 216(6):1715–1730

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Xu H, Ren D (2015) Lysosomal physiology. Annu Rev Physiol 77:57–80

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Yudowski GA, Puthenveedu MA, Henry AG, von Zastrow M (2009) Cargo-mediated regulation of a rapid Rab4-dependent recycling pathway. Mol Biol Cell 20(11):2774–2784

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang X, Li X, Xu H (2012) Phosphoinositide isoforms determine compartment-specific ion channel activity. Proc Natl Acad Sci U S A 109(28):11384–11389

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhong XZ, Dong XP (2015) Lysosome electrophysiology. Methods Cell Biol 126:197–215

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by funding from the National Science and Technology Council (MOST110-2320-B002-022-MY3, National Health Research Institutes (NHRI-EX112-11119SC), and National Taiwan University (112L7818) to C.-C. C.

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Correspondence to Cheng-Chang Chen .

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Chen, CC. (2023). Electrophysiological Techniques on the Study of Endolysosomal Ion Channels. In: Wahl-Schott, C., Biel, M. (eds) Endolysosomal Voltage-Dependent Cation Channels. Handbook of Experimental Pharmacology, vol 278. Springer, Cham. https://doi.org/10.1007/164_2023_638

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