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BetaPix up-regulates Na+/H+ exchanger 3 through a Shank2-mediated protein-protein interaction

 Won-Joo Kim  ;  Ji Hyun Lee  ;  Jihyun Yi  ;  Yang-Je Cho  ;  Kyoung Heo  ;  Sung Hee Lee  ;  So Won Kim  ;  Myeong-Kyu Kim  ;  Kyung Hwan Kim  ;  Byung In Lee  ;  Min Goo Lee 
 JOURNAL OF BIOLOGICAL CHEMISTRY, Vol.285(11) : 8104-8113, 2010 
Journal Title
Issue Date
Animals ; Cell Polarity/physiology ; Cells, Cultured ; Cricetinae ; Endothelial Cells/cytology ; Endothelial Cells/enzymology* ; Fibroblasts/cytology ; Fibroblasts/enzymology ; Guanine Nucleotide Exchange Factors/metabolism* ; Humans ; Kidney/cytology ; Membrane Proteins/metabolism ; Nerve Tissue Proteins/metabolism* ; Protein Binding/physiology ; Protons ; Rats ; Rats, Sprague-Dawley ; Rho Guanine Nucleotide Exchange Factors ; Sodium/metabolism ; Sodium-Hydrogen Exchanger 3 ; Sodium-Hydrogen Exchangers/metabolism* ; Up-Regulation/physiology ; rho GTP-Binding Proteins/metabolism
Na(+)/H(+) exchanger 3 (NHE3) plays an important role in neutral Na(+) transport in mammalian epithelial cells. The Rho family of small GTPases and the PDZ (PSD-95/discs large/ZO-1) domain-based adaptor Shank2 are known to regulate the membrane expression and activity of NHE3. In this study we examined the role of betaPix, a guanine nucleotide exchange factor for the Rho GTPase and a strong binding partner to Shank2, in NHE3 regulation using integrated molecular and physiological approaches. Immunoprecipitation and pulldown assays revealed that NHE3, Shank2, and betaPix form a macromolecular complex when expressed heterologously in mammalian cells as well as endogenously in rat colon, kidney, and pancreas. In addition, these proteins co-segregated at the apical surface of rat colonic epithelial cells, as detected by immunofluorescence staining. When expressed in PS120/NHE3 cells, betaPix increased membrane expression and basal activity of NHE3. Interestingly, the effects of betaPix on NHE3 were abolished by cotransfection with dominant-negative Shank2 mutants and by treatment with Clostridium difficile toxin B, a Rho GTPase inhibitor, indicating that Shank2 and Rho GTPases are involved in betaPix-mediated NHE3 regulation. Knockdown of endogenous betaPix by RNA interference decreased Shank2-induced increase of NHE3 membrane expression in HEK 293T cells. These results indicate that betaPix up-regulates NHE3 membrane expression and activity by Shank2-mediated protein-protein interaction and by activating Rho GTPases in the apical regions of epithelial cells.
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1. College of Medicine (의과대학) > Dept. of Pharmacology (약리학교실) > 1. Journal Papers
Yonsei Authors
Kim, Kyung Hwan(김경환)
Kim, Joo Young(김주영) ORCID logo https://orcid.org/0000-0003-2623-1491
Park, Hyun Woo(박현우)
Lee, Min Goo(이민구) ORCID logo https://orcid.org/0000-0001-7436-012X
Lee, Jung Soo(이정수)
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