Bradykinin B2 receptors activate Na+/H+ exchange in mIMCD-3 cells via Janus kinase 2 and Ca2+/calmodulin.
We used a cultured murine cell model of the inner medullary collecting duct (mIMCD-3 cells) to examine the regulation of the ubiquitous sodium-proton exchanger, Na+/H+ exchanger isoform 1 (NHE-1), by a prototypical G protein-coupled receptor, the bradykinin B2 receptor. Bradykinin rapidly activates NHE-1 in a concentration-dependent manner as assessed by proton microphysiometry of quiescent cells and by 2'-7'-bis[2-carboxymethyl]-5(6)-carboxyfluorescein fluorescence measuring the accelerated rate of pH(i) recovery from an imposed acid load. The activation of NHE-1 is blocked by inhibitors of the bradykinin B2 receptor, phospholipase C, Ca2+/calmodulin (CaM), and Janus kinase 2 (Jak2), but not by pertussis toxin or by inhibitors of protein kinase C and phosphatidylinositol 3'-kinase. Immunoprecipitation studies showed that bradykinin stimulates the assembly of a signal transduction complex that includes CaM, Jak2, and NHE-1. CaM appears to be a direct substrate for phosphorylation by Jak2 as measured by an in vitro kinase assay. We propose that Jak2 is a new indirect regulator of NHE-1 activity, which modulates the activity of NHE-1 by increasing the tyrosine phosphorylation of CaM and most likely by increasing the binding of CaM to NHE-1.
Medical and Research Services of the Ralph H. Johnson Veterans Affairs Medical Center, and Department of Medicine of the Medical University of South Carolina, Charleston, South Carolina 29425, USA.
Y V
YV
Mukhin
T
T
Vlasova
A A
AA
Jaffa
G
G
Collinsworth
J L
JL
Bell
B G
BG
Tholanikunnel
T
T
Pettus
W
W
Fitzgibbon
D W
DW
Ploth
J R
JR
Raymond
M N
MN
Garnovskaya
D000818:Animals; D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D053614:Janus Kinase 2; D007679:Kidney Medulla / Q000166:cytology / Q000502:physiology; D007700:Kinetics; D051379:Mice; D008822:Mice, Transgenic; D008954:Models, Biological; D037342:Pertussis Toxin; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins; D043782:Receptor, Bradykinin B2; D018002:Receptors, Bradykinin / Q000235:genetics / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology; D013539:Simian virus 40 / Q000235:genetics; D012964:Sodium / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*; D014158:Transcription, Genetic; D010738:Type C Phospholipases / Q000378:metabolism; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D002147:Calmodulin; D004791:Enzyme Inhibitors; D011518:Proto-Oncogene Proteins; D043782:Receptor, Bradykinin B2; D018002:Receptors, Bradykinin; D017923:Sodium-Hydrogen Exchangers; D010566:Virulence Factors, Bordetella; C069139:growth factor-activatable Na-H exchanger NHE-1; D012964:Sodium; D037342:Pertussis Toxin; D011505:Protein-Tyrosine Kinases; C507926:Jak2 protein, mouse; D053614:Janus Kinase 2; D010738:Type C Phospholipases; D001920:Bradykinin; D002118:Calcium