title: Spatial control of proton pump H,K-ATPase docking at the apical membrane by phosphorylation-coupled ezrin-syntaxin 3 interaction.
abstract: The digestive function of the stomach depends on acidification of the gastric lumen. Acid secretion into the lumen is triggered by activation of a cAMP-dependent protein kinase (PKA) cascade, which ultimately results in the insertion of gastric H,K-ATPases into the apical plasma membranes of parietal cells. A coupling protein is ezrin whose phosphorylation at Ser-66 by PKA is required for parietal cell activation. However, little is known regarding the molecular mechanism(s) by which ezrin operates in gastric acid secretion. Here we show that phosphorylation of Ser-66 induces a conformational change of ezrin that enables its association with syntaxin 3 (Stx3) and provides a spatial cue for H,K-ATPase trafficking. This conformation-dependent association is specific for Stx3, and the binding interface is mapped to the N-terminal region. Biochemical analyses show that inhibition of ezrin phosphorylation at Ser-66 prevents ezrin-Stx3 association and insertion of H,K-ATPase into the apical plasma membrane of parietal cells. Using atomic force microscopic analyses, our study revealed that phosphorylation of Ser-66 induces unfolding of ezrin molecule to allow Stx3 binding to its N terminus. Given the essential role of Stx3 in polarized secretion, our study presents the first evidence in which phosphorylation-induced conformational rearrangement of the ezrin molecule provides a spatial cue for polarized membrane trafficking in epithelial cells.
authors.affiliation: From the Anhui Key Laboratory for Cellular Dynamics and Chemical Biology, University of Science and Technology of China School of Life Science, Hefei, China 230027.
authors.forename: Huijuan
authors.initials: H
authors.lastname: Yu
authors.affiliation: From the Anhui Key Laboratory for Cellular Dynamics and Chemical Biology, University of Science and Technology of China School of Life Science, Hefei, China 230027, Morehouse School of Medicine, Atlanta, Georgia 30310, and.
authors.forename: Jiajia
authors.initials: J
authors.lastname: Zhou
authors.affiliation: Laboratory of Plasma Membrane, Graduate School of Biostudies, Kyoto University, Kyoto 606-8502, Japan.
authors.forename: Hirohide
authors.initials: H
authors.lastname: Takahashi
authors.affiliation: Morehouse School of Medicine, Atlanta, Georgia 30310, and.
authors.forename: William
authors.initials: W
authors.lastname: Yao
authors.affiliation: Laboratory of Plasma Membrane, Graduate School of Biostudies, Kyoto University, Kyoto 606-8502, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Suzuki
authors.affiliation: From the Anhui Key Laboratory for Cellular Dynamics and Chemical Biology, University of Science and Technology of China School of Life Science, Hefei, China 230027.
authors.forename: Xiao
authors.initials: X
authors.lastname: Yuan
authors.affiliation: Laboratory of Plasma Membrane, Graduate School of Biostudies, Kyoto University, Kyoto 606-8502, Japan.
authors.forename: Shige H
authors.initials: SH
authors.lastname: Yoshimura
authors.affiliation: From the Anhui Key Laboratory for Cellular Dynamics and Chemical Biology, University of Science and Technology of China School of Life Science, Hefei, China 230027, Graduate School, Beijing University of Chinese Medicine, Beijing 100086, China.
authors.forename: Yin
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: From the Anhui Key Laboratory for Cellular Dynamics and Chemical Biology, University of Science and Technology of China School of Life Science, Hefei, China 230027.
authors.forename: Ya
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Morehouse School of Medicine, Atlanta, Georgia 30310, and.
authors.forename: Nerimiah
authors.initials: N
authors.lastname: Emmett
authors.affiliation: Morehouse School of Medicine, Atlanta, Georgia 30310, and.
authors.forename: Vincent
authors.initials: V
authors.lastname: Bond
authors.affiliation: From the Anhui Key Laboratory for Cellular Dynamics and Chemical Biology, University of Science and Technology of China School of Life Science, Hefei, China 230027.
authors.forename: Dongmei
authors.initials: D
authors.lastname: Wang
authors.affiliation: Graduate School, Beijing University of Chinese Medicine, Beijing 100086, China.
authors.forename: Xia
authors.initials: X
authors.lastname: Ding
authors.affiliation: Laboratory of Plasma Membrane, Graduate School of Biostudies, Kyoto University, Kyoto 606-8502, Japan, takeyasu@kyoto.ac.jp.
authors.forename: Kunio
authors.initials: K
authors.lastname: Takeyasu
authors.affiliation: From the Anhui Key Laboratory for Cellular Dynamics and Chemical Biology, University of Science and Technology of China School of Life Science, Hefei, China 230027, yaoxb@ustc.edu.cn.
authors.forename: Xuebiao
authors.initials: X
authors.lastname: Yao
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D017506:H(+)-K(+)-Exchanging ATPase / Q000378:metabolism*; D010295:Parietal Cells, Gastric / Q000166:cytology / Q000378:metabolism*; D010766:Phosphorylation / Q000502:physiology; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology; D050765:Qa-SNARE Proteins / Q000378:metabolism*; D011817:Rabbits; D000099149:Ezrin
chemical_list: D003598:Cytoskeletal Proteins; D017506:H(+)-K(+)-Exchanging ATPase; D050765:Qa-SNARE Proteins; D000099149:Ezrin
keywords: A-kinase Anchoring Protein (AKAP); ABC Transporter; ATPase; Atomic Force Microscopy (AFM); Cell Polarity; Cytoskeleton; Epithelial Cell; Exocytosis; Ezrin; H+-ATPase