Spatial Control of Proton Pump H,K-ATPase Docking at the Apical Membrane by Phosphorylation-coupled Ezrin-Syntaxin 3 Interaction

• Published in: The Journal of biological chemistry Vol. 289; no. 48; pp. 33333 - 33342
• Main Authors: Yu, Huijuan, Zhou, Jiajia, Takahashi, Hirohide, Yao, William, Suzuki, Yuki, Yuan, Xiao, Yoshimura, Shige H., Zhang, Yin, Liu, Ya, Emmett, Nerimiah, Bond, Vincent, Wang, Dongmei, Ding, Xia, Takeyasu, Kunio, Yao, Xuebiao
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 28-11-2014; American Society for Biochemistry and Molecular Biology
• Subjects: A-kinase Anchoring Protein (AKAP); ABC Transporter; Animals; Atomic Force Microscopy (AFM); ATPase; Cell Biology; Cell Membrane - metabolism; Cell Polarity; Cells, Cultured; Cytoskeletal Proteins - metabolism; Cytoskeleton; Epithelial Cell; Exocytosis; Ezrin; H(+)-K(+)-Exchanging ATPase - metabolism; H+-ATPase; Parietal Cells, Gastric - cytology; Parietal Cells, Gastric - metabolism; Phosphorylation - physiology; Protein Structure, Tertiary; Protein Transport - physiology; Qa-SNARE Proteins - metabolism; Rabbits; Cell Polarity; Atomic Force Microscopy (AFM); ABC Transporter; ATPase; Ezrin; Epithelial Cell; Cytoskeleton; H+-ATPase; A-kinase Anchoring Protein (AKAP); Exocytosis
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M114.581280

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.
Background: Polarized acid secretion in gastric parietal cells requires ezrin and its phosphorylation at Ser-66.
Results: Phosphorylation of Ser-66 induces ezrin conformational change, which enables ezrin to interact with syntaxin 3.
Conclusion: Conformational change of ezrin provides a spatial cue for apical trafficking of H,K-ATPase.
Significance: Ezrin conformation orchestrates the polarized vesicle trafficking in epithelial cells.