Lysophosphatidic acid induces exocytic trafficking of Na(+)/H(+) exchanger 3 by E3KARP-dependent activation of phospholipase C.
Lysophosphatidic acid (LPA) stimulates Na(+)/H(+) exchanger 3 (NHE3) activity in opossum kidney proximal tubule (OK) cells by increasing the apical membrane amount of NHE3. This occurs by stimulation of exocytic trafficking of NHE3 to the apical plasma membrane by an E3KARP-dependent mechanism. However, it is still unclear how E3KARP leads to the LPA-induced exocytosis of NHE3. In the current study, we demonstrate that stable expression of exogenous E3KARP increases LPA-induced phospholipase C (PLC) activation and subsequent elevation of intracellular Ca(2+) in opossum kidney proximal tubule (OK) cells. Pretreatment with U73122, a PLC inhibitor, prevented the LPA-induced NHE3 activation and the exocytic trafficking of NHE3. To understand how the elevation of intracellular Ca(2+) leads to the stimulation of NHE3, we pretreated OK cells with BAPTA-AM, an intracellular Ca(2+) chelator. BAPTA-AM completely blocked the LPA-induced increase of NHE3 activity and surface NHE3 amount by decreasing the LPA-induced exocytic trafficking of NHE3. Pretreatment with GF109203X, a PKC inhibitor, did not affect the percent of LPA-induced NHE3 activation and increase of surface NHE3 amount. From these results, we suggest that E3KARP plays a necessary role in LPA-induced PLC activation, and that PLC-dependent elevation of intracellular Ca(2+) but not PKC activation is necessary for the LPA-induced increase of NHE3 exocytosis.
Choi
Jung Woong
JW
Department of Physiology and Medicine, Gastrointestinal Division, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Lee-Kwon
Whaseon
W
Jeon
Eun Su
ES
Kang
Yong Jung
YJ
Kawano
Kazuya
K
Kim
Hyeon Soo
HS
Suh
Pann-Ghill
PG
Donowitz
Mark
M
Kim
Jae Ho
JH
D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D006224:Cricetinae; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005089:Exocytosis / Q000187:drug effects*; D007211:Indoles / Q000494:pharmacology; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000378:metabolism; D008246:Lysophospholipids / Q000494:pharmacology*; D008301:Maleimides / Q000494:pharmacology; D009893:Opossums / Q000502:physiology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D021381:Protein Transport; D015398:Signal Transduction; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism*
D002614:Chelating Agents; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors; D007211:Indoles; D008246:Lysophospholipids; D008301:Maleimides; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D011493:Protein Kinase C; D010738:Type C Phospholipases; C070515:bisindolylmaleimide I; C032881:lysophosphatidic acid; D002118:Calcium