Regulation of the epithelial Na+ channel by the protein kinase CK2.
CK2 is a ubiquitous, pleiotropic, and constitutively active Ser/Thr protein kinase that controls protein expression, cell signaling, and ion channel activity. Phosphorylation sites for CK2 are located in the C terminus of both beta- and gamma-subunits of the epithelial Na(+) channel (ENaC). We examined the role of CK2 on the regulation of both endogenous ENaC in native murine epithelia and in Xenopus oocytes expressing rENaC. In Ussing chamber experiments with mouse airways, colon, and cultured M1-collecting duct cells, amiloride-sensitive Na(+) transport was inhibited dose-dependently by the selective CK2 inhibitor 4,5,6,7-tetrabromobenzotriazole (TBB). In oocytes, ENaC currents were also inhibited by TBB and by the structurally unrelated inhibitors heparin and poly(E:Y). Expression of a trimeric channel lacking both CK2 sites (alphabeta(S631A)gamma(T599A)) produced a largely attenuated amiloride-sensitive whole cell conductance and rendered the mutant channel insensitive to CK2. In Xenopus oocytes, CK2 was translocated to the cell membrane upon expression of wt-ENaC but not of alphabeta(S631A)gamma(T599A)-ENaC. Phosphorylation by CK2 is essential for ENaC activation, and to a lesser degree, it also controls membrane expression of alphabetagamma-ENaC. Channels lacking the Nedd4-2 binding motif in beta-ENaC (R561X, Y618A) no longer required the CK2 site for channel activity and siRNA-knockdown of Nedd4-2 eliminated the effects of TBB. This implies a role for CK2 in inhibiting the Nedd4-2 pathway. We propose that the C terminus of beta-ENaC is targeted by this essential, conserved pleiotropic kinase that directs its constitutive activity toward many cellular protein complexes.
Bachhuber
Tanja
T
Institut für Physiologie, Universität Regensburg, Universitätsstrasse 31, D-93053 Regensburg, Germany.
Almaça
Joana
J
Aldehni
Fadi
F
Mehta
Anil
A
Amaral
Margarida D
MD
Schreiber
Rainer
R
Kunzelmann
Karl
K
D000818:Animals; D047390:Casein Kinase II / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D004594:Electrophysiology; D004789:Enzyme Activation; D004847:Epithelial Cells / Q000187:drug effects / Q000201:enzymology; D053503:Epithelial Sodium Channels / Q000235:genetics / Q000378:metabolism*; D005260:Female; D051379:Mice; D008810:Mice, Inbred C57BL; D009865:Oocytes; D018408:Patch-Clamp Techniques; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D021381:Protein Transport; D051381:Rats; D012964:Sodium / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D014230:Triazoles / Q000494:pharmacology; D014982:Xenopus laevis
C405354:4,5,6,7-tetrabromobenzotriazole; D053503:Epithelial Sodium Channels; D047428:Protein Kinase Inhibitors; D014230:Triazoles; D012964:Sodium; D047390:Casein Kinase II