PIKfyve upregulates CFTR activity.
The cystic fibrosis transmembrane conductance regulator (CFTR) is a cAMP-activated Cl(-) channel critically important in Cl(-) secreting epithelia. Mutations in the CFTR gene, such as (DeltaF508)CFTR leads to cystic fibrosis, a severe disease with defective Cl(-) secretion. CFTR is stimulated by the serum and glucocorticoid-inducible kinase SGK1. The SGK1 dependent regulation of several carriers and channels involves the phosphatidylinositol-3-phosphate-5-kinase PIKfyve, which similarly mediates the regulation of glucose carriers by PKB/Akt. The present study was thus performed to elucidate whether PKB/Akt and PIKfyve are regulators of CFTR. To this end CFTR or (DeltaF508)CFTR were expressed in Xenopus oocytes alone or together with PKB, PIKfyve or the SGK1/PKB resistant mutant (S318A)PIKfyve, and the current generated by cAMP upregulation with 10muM forskolin+1mM IBMX determined utilizing dual electrode voltage clamp. As a result, forskolin/IBMX treatment triggered a current (I(cAMP)) in CFTR-expressing Xenopus oocytes, but not in oocytes expressing (DeltaF508)CFTR. Coexpression of PKB/Akt and PIKfyve, but not of (S318A)PIKfyve, stimulated I(cAMP) in CFTR-expressing ( approximately 2- to 3-fold) but not in (DeltaF508)CFTR-expressing or water injected Xenopus oocytes. Immunohistochemistry revealed that the coexpression of PIKfyve, but not of (S318A)PIKfyve, enhanced the CFTR protein abundance but not the (DeltaF508)CFTR protein abundance in CFTR or (DeltaF508)CFTR-expressing oocytes. The present observations reveal a novel powerful regulator of intact but not of defective CFTR.
Department of Physiology, University of Tübingen, Gmelinstrasse 5, D-72076 Tübingen, Germany.
Eva-Maria
EM
Gehring
Rebecca S
RS
Lam
Gulab
G
Siraskar
Evgenia
E
Koutsouki
Guiscard
G
Seebohm
Oana N
ON
Ureche
Liviu
L
Ureche
Ravshan
R
Baltaev
Jeremy M
JM
Tavare
Florian
F
Lang
D064413:3-Phosphoinositide-Dependent Protein Kinases; D000818:Animals; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D009865:Oocytes; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D015854:Up-Regulation; D014982:Xenopus laevis
D064413:3-Phosphoinositide-Dependent Protein Kinases; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; C505032:CFTR protein, human; C492139:PIKFYVE protein, human