title: Transport through recycling endosomes requires EHD1 recruitment by a phosphatidylserine translocase.
abstract: P4-ATPases translocate aminophospholipids, such as phosphatidylserine (PS), to the cytosolic leaflet of membranes. PS is highly enriched in recycling endosomes (REs) and is essential for endosomal membrane traffic. Here, we show that PS flipping by an RE-localized P4-ATPase is required for the recruitment of the membrane fission protein EHD1. Depletion of ATP8A1 impaired the asymmetric transbilayer distribution of PS in REs, dissociated EHD1 from REs, and generated aberrant endosomal tubules that appear resistant to fission. EHD1 did not show membrane localization in cells defective in PS synthesis. ATP8A2, a tissue-specific ATP8A1 paralogue, is associated with a neurodegenerative disease (CAMRQ). ATP8A2, but not the disease-causative ATP8A2 mutant, rescued the endosomal defects in ATP8A1-depleted cells. Primary neurons from Atp8a2-/- mice showed a reduced level of transferrin receptors at the cell surface compared to Atp8a2+/+ mice. These findings demonstrate the role of P4-ATPase in membrane fission and give insight into the molecular basis of CAMRQ.
authors.affiliation: Department of Health Chemistry, Graduate School of Pharmaceutical Sciences University of Tokyo, Tokyo, Japan.
authors.forename: Shoken
authors.initials: S
authors.lastname: Lee
authors.affiliation: Department of Health Chemistry, Graduate School of Pharmaceutical Sciences University of Tokyo, Tokyo, Japan.
authors.forename: Yasunori
authors.initials: Y
authors.lastname: Uchida
authors.affiliation: Departments of Biochemistry and Molecular Biology and Ophthalmology and Visual Sciences, Centre for Macular Research University of British Columbia, Vancouver, BC, Canada.
authors.forename: Jiao
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Health Chemistry, Graduate School of Pharmaceutical Sciences University of Tokyo, Tokyo, Japan.
authors.forename: Tatsuyuki
authors.initials: T
authors.lastname: Matsudaira
authors.affiliation: Department of Pharmacology, Osaka Medical College, Takatsuki-city Osaka, Japan.
authors.forename: Takatoshi
authors.initials: T
authors.lastname: Nakagawa
authors.affiliation: Lipid Biology Laboratory, RIKEN, Wako-shi Saitama, Japan.
authors.forename: Takuma
authors.initials: T
authors.lastname: Kishimoto
authors.affiliation: Department of Health Chemistry, Graduate School of Pharmaceutical Sciences University of Tokyo, Tokyo, Japan Lipid Biology Laboratory, RIKEN, Wako-shi Saitama, Japan.
authors.forename: Kojiro
authors.initials: K
authors.lastname: Mukai
authors.affiliation: Lipid Biology Laboratory, RIKEN, Wako-shi Saitama, Japan.
authors.forename: Takehiko
authors.initials: T
authors.lastname: Inaba
authors.affiliation: Lipid Biology Laboratory, RIKEN, Wako-shi Saitama, Japan.
authors.forename: Toshihide
authors.initials: T
authors.lastname: Kobayashi
authors.affiliation: Departments of Biochemistry and Molecular Biology and Ophthalmology and Visual Sciences, Centre for Macular Research University of British Columbia, Vancouver, BC, Canada.
authors.forename: Robert S
authors.initials: RS
authors.lastname: Molday
authors.affiliation: Department of Health Chemistry, Graduate School of Pharmaceutical Sciences University of Tokyo, Tokyo, Japan Pathological Cell Biology Laboratory, Graduate School of Pharmaceutical Sciences University of Tokyo, Tokyo, Japan tom_taguchi@mol.f.u-tokyo.ac.jp harai@mol.f.u-tokyo.ac.jp.
authors.forename: Tomohiko
authors.initials: T
authors.lastname: Taguchi
authors.affiliation: Department of Health Chemistry, Graduate School of Pharmaceutical Sciences University of Tokyo, Tokyo, Japan Pathological Cell Biology Laboratory, Graduate School of Pharmaceutical Sciences University of Tokyo, Tokyo, Japan tom_taguchi@mol.f.u-tokyo.ac.jp harai@mol.f.u-tokyo.ac.jp.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Arai
mesh_terms: D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D000704:Analysis of Variance; D000818:Animals; D001426:Bacterial Proteins; D001692:Biological Transport / Q000502:physiology; D015153:Blotting, Western; D019556:COS Cells; D002522:Chlorocebus aethiops; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D011992:Endosomes / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D008954:Models, Biological; D010718:Phosphatidylserines / Q000378:metabolism; D049294:Phospholipid Transfer Proteins / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D034622:RNA Interference; D013301:Streptolysins; D033921:Vesicular Transport Proteins / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; D018076:DNA, Complementary; C120700:EHD1 protein, human; D010718:Phosphatidylserines; D049294:Phospholipid Transfer Proteins; D013301:Streptolysins; D033921:Vesicular Transport Proteins; C025964:streptolysin O; C580722:ATP8A1 protein, human; C545446:ATP8a2 protein, mouse; D000251:Adenosine Triphosphatases
keywords: ATP8A1; EHD1; phosphatidylserine; phospholipid flippase; recycling endosomes