title: Calcium release through P2X4 activates calmodulin to promote endolysosomal membrane fusion.
abstract: Intra-endolysosomal Ca(2+) release is required for endolysosomal membrane fusion with intracellular organelles. However, the molecular mechanisms for intra-endolysosomal Ca(2+) release and the downstream Ca(2+) targets involved in the fusion remain elusive. Previously, we demonstrated that endolysosomal P2X4 forms channels activated by luminal adenosine triphosphate in a pH-dependent manner. In this paper, we show that overexpression of P2X4, as well as increasing endolysosomal P2X4 activity by alkalinization of endolysosome lumen, promoted vacuole enlargement in cells and endolysosome fusion in a cell-free assay. These effects were prevented by inhibiting P2X4, expressing a dominant-negative P2X4 mutant, and disrupting the P2X4 gene. We further show that P2X4 and calmodulin (CaM) form a complex at endolysosomal membrane where P2X4 activation recruits CaM to promote fusion and vacuolation in a Ca(2+)-dependent fashion. Moreover, P2X4 activation-triggered fusion and vacuolation were suppressed by inhibiting CaM. Our data thus suggest a new molecular mechanism for endolysosomal membrane fusion involving P2X4-mediated endolysosomal Ca(2+) release and subsequent CaM activation.
authors.affiliation: Department of Physiology and Biophysics, Dalhousie University, Halifax B3H 4R2, Nova Scotia, Canada.
authors.forename: Qi
authors.initials: Q
authors.lastname: Cao
authors.affiliation: Department of Physiology and Biophysics, Dalhousie University, Halifax B3H 4R2, Nova Scotia, Canada.
authors.forename: Xi Zoë
authors.initials: XZ
authors.lastname: Zhong
authors.affiliation: Department of Physiology and Biophysics, Dalhousie University, Halifax B3H 4R2, Nova Scotia, Canada.
authors.forename: Yuanjie
authors.initials: Y
authors.lastname: Zou
authors.affiliation: Department of Pharmacology, University of Cambridge, Cambridge CB2 1PD, England, UK.
authors.forename: Ruth
authors.initials: R
authors.lastname: Murrell-Lagnado
authors.affiliation: Department of Integrative Biology and Pharmacology, The University of Texas Health Science Center at Houston, Houston, TX 77030.
authors.forename: Michael X
authors.initials: MX
authors.lastname: Zhu
authors.affiliation: Department of Physiology and Biophysics, Dalhousie University, Halifax B3H 4R2, Nova Scotia, Canada xpdong@dal.ca.
authors.forename: Xian-Ping
authors.initials: XP
authors.lastname: Dong
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001692:Biological Transport; D019556:COS Cells; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002460:Cell Line; D002522:Chlorocebus aethiops; D057809:HEK293 Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D008561:Membrane Fusion / Q000502:physiology*; D051379:Mice; D058848:Receptors, Purinergic P2X4 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
chemical_list: D002147:Calmodulin; D058848:Receptors, Purinergic P2X4; D000255:Adenosine Triphosphate; D002118:Calcium