title: The Two-pore channel (TPC) interactome unmasks isoform-specific roles for TPCs in endolysosomal morphology and cell pigmentation.
abstract: The two-pore channels (TPC1 and TPC2) belong to an ancient family of intracellular ion channels expressed in the endolysosomal system. Little is known about how regulatory inputs converge to modulate TPC activity, and proposed activation mechanisms are controversial. Here, we compiled a proteomic characterization of the human TPC interactome, which revealed that TPCs complex with many proteins involved in Ca(2+) homeostasis, trafficking, and membrane organization. Among these interactors, TPCs were resolved to scaffold Rab GTPases and regulate endomembrane dynamics in an isoform-specific manner. TPC2, but not TPC1, caused a proliferation of endolysosomal structures, dysregulating intracellular trafficking, and cellular pigmentation. These outcomes required both TPC2 and Rab activity, as well as their interactivity, because TPC2 mutants that were inactive, or rerouted away from their endogenous expression locale, or deficient in Rab binding, failed to replicate these outcomes. Nicotinic acid adenine dinucleotide phosphate (NAADP)-evoked Ca(2+) release was also impaired using either a Rab binding-defective TPC2 mutant or a Rab inhibitor. These data suggest a fundamental role for the ancient TPC complex in trafficking that holds relevance for lysosomal proliferative scenarios observed in disease.
authors.affiliation: Department of Pharmacology, University of Minnesota, Minneapolis, MN 55455;
authors.forename: Yaping
authors.initials: Y
authors.lastname: Lin-Moshier
authors.affiliation: Department of Pharmacology, University of Minnesota, Minneapolis, MN 55455;
authors.forename: Michael V
authors.initials: MV
authors.lastname: Keebler
authors.affiliation: Department of Cell and Developmental Biology, University College London, London WC1E 6BT, United Kingdom; and.
authors.forename: Robert
authors.initials: R
authors.lastname: Hooper
authors.affiliation: Department of Pharmacology, University of Minnesota, Minneapolis, MN 55455;
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Boulware
authors.affiliation: Department of Pharmacology, University of Minnesota, Minneapolis, MN 55455;
authors.forename: Xiaolong
authors.initials: X
authors.lastname: Liu
authors.affiliation: Department of Cell and Developmental Biology, University College London, London WC1E 6BT, United Kingdom; and.
authors.forename: Dev
authors.initials: D
authors.lastname: Churamani
authors.affiliation: Departments of Anatomy, Cell Biology, and Pharmacology and the Center for Substance Abuse, Temple University School of Medicine, Philadelphia, PA 19140.
authors.forename: Mary E
authors.initials: ME
authors.lastname: Abood
authors.affiliation: Department of Pharmacology, University of Minnesota, Minneapolis, MN 55455;
authors.forename: Timothy F
authors.initials: TF
authors.lastname: Walseth
authors.affiliation: Departments of Anatomy, Cell Biology, and Pharmacology and the Center for Substance Abuse, Temple University School of Medicine, Philadelphia, PA 19140.
authors.forename: Eugen
authors.initials: E
authors.lastname: Brailoiu
authors.affiliation: Department of Cell and Developmental Biology, University College London, London WC1E 6BT, United Kingdom; and.
authors.forename: Sandip
authors.initials: S
authors.lastname: Patel
authors.affiliation: Department of Pharmacology, University of Minnesota, Minneapolis, MN 55455; march029@umn.edu.
authors.forename: Jonathan S
authors.initials: JS
authors.lastname: Marchant
mesh_terms: D000818:Animals; D015220:Calcium Channels / Q000378:metabolism*; D020013:Calcium Signaling; D049109:Cell Proliferation; D002846:Chromatography, Affinity; D011992:Endosomes / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D009249:NADP / Q000031:analogs & derivatives / Q000378:metabolism; D010858:Pigmentation; D011485:Protein Binding; D020033:Protein Isoforms / Q000378:metabolism; D015203:Reproducibility of Results; D014981:Xenopus; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000097903:Two-Pore Channels
chemical_list: D015220:Calcium Channels; D009249:NADP; D020033:Protein Isoforms; D020691:rab GTP-Binding Proteins; C407357:TPCN1 protein, human; D000097903:Two-Pore Channels; C530224:TPCN2 protein, human; C024376:NAADP
keywords: Ca2+ signaling; Xenopus; lysosome