cGMP inhibition of Na+/H+ antiporter 3 (NHE3) requires PDZ domain adapter NHERF2, a broad specificity protein kinase G-anchoring protein.
Electroneutral NaCl absorption mediated by Na+/H+ exchanger 3 (NHE3) is important in intestinal and renal functions related to water/Na+ homeostasis. cGMP inhibits NHE3 in intact epithelia. However, unexpectedly it failed to inhibit NHE3 stably transfected in PS120 cells, even upon co-expression of cGMP-dependent protein kinase type II (cGKII). Additional co-expression of NHERF2, the tandem PDZ domain adapter protein involved in cAMP inhibition of NHE3, restored cGMP as well as cAMP inhibition, whereas NHERF1 solely restored cAMP inhibition. In vitro conditions were identified in which NHERF2 but not NHERF1 bound cGKII. The NHERF2 PDZ2 C terminus, which binds NHE3, also bound cGKII. A non-myristoylated mutant of cGKII did not support cGMP inhibition of NHE3. Although cGKI also bound NHERF2 in vitro, it did not evoke inhibition of NHE3 unless a myristoylation site was added. These results show that NHERF2, acting as a novel protein kinase G-anchoring protein, is required for cGMP inhibition of NHE3 and that cGKII must be bound both to the plasma membrane by its myristoyl anchor and to NHERF2 to inhibit NHE3.
Cha
Boyoung
B
Department of Physiology, GI Division, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Kim
Jae Ho
JH
Hut
Hans
H
Hogema
Boris M
BM
Nadarja
Janani
J
Zizak
Mirza
M
Cavet
Megan
M
Lee-Kwon
Whaseon
W
Lohmann
Suzanne M
SM
Smolenski
Albert
A
Tse
Chung Ming
CM
Yun
Chris
C
de Jonge
Hugo R
HR
Donowitz
Mark
M
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism*; D062967:Cyclic GMP-Dependent Protein Kinase Type II; D017869:Cyclic GMP-Dependent Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000737:chemistry* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000737:chemistry; D006801:Humans; D047468:Immunoprecipitation; D046911:Macromolecular Substances / Q000378:metabolism; D009154:Mutation; D019814:Myristic Acid / Q000737:chemistry; D009893:Opossums; D010750:Phosphoproteins; D011485:Protein Binding; D011494:Protein Kinases / Q000737:chemistry; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors* / Q000502:physiology*; D013379:Substrate Specificity; D014162:Transfection
D003598:Cytoskeletal Proteins; D046911:Macromolecular Substances; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C000613385:SLC9A3 protein, human; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; C093376:sodium-hydrogen exchanger regulatory factor; D019814:Myristic Acid; D000242:Cyclic AMP; D011494:Protein Kinases; D062967:Cyclic GMP-Dependent Protein Kinase Type II; D017869:Cyclic GMP-Dependent Protein Kinases; C569729:PRKG2 protein, human; D019204:GTP-Binding Proteins; D006152:Cyclic GMP