Interaction with 14-3-3 proteins promotes functional expression of the potassium channels TASK-1 and TASK-3.
The two-pore-domain potassium channels TASK-1, TASK-3 and TASK-5 possess a conserved C-terminal motif of five amino acids. Truncation of the C-terminus of TASK-1 strongly reduced the currents measured after heterologous expression in Xenopus oocytes or HEK293 cells and decreased surface membrane expression of GFP-tagged channel proteins. Two-hybrid analysis showed that the C-terminal domain of TASK-1, TASK-3 and TASK-5, but not TASK-4, interacts with isoforms of the adapter protein 14-3-3. A pentapeptide motif at the extreme C-terminus of TASK-1, RRx(S/T)x, was found to be sufficient for weak but significant interaction with 14-3-3, whereas the last 40 amino acids of TASK-1 were required for strong binding. Deletion of a single amino acid at the C-terminal end of TASK-1 or TASK-3 abolished binding of 14-3-3 and strongly reduced the macroscopic currents observed in Xenopus oocytes. TASK-1 mutants that failed to interact with 14-3-3 isoforms (V411*, S410A, S410D) also produced only very weak macroscopic currents. In contrast, the mutant TASK-1 S409A, which interacts with 14-3-3-like wild-type channels, displayed normal macroscopic currents. Co-injection of 14-3-3zeta cRNA increased TASK-1 current in Xenopus oocytes by about 70 %. After co-transfection in HEK293 cells, TASK-1 and 14-3-3zeta (but not TASK-1DeltaC5 and 14-3-3zeta) could be co-immunoprecipitated. Furthermore, TASK-1 and 14-3-3 could be co-immunoprecipitated in synaptic membrane extracts and postsynaptic density membranes. Our findings suggest that interaction of 14-3-3 with TASK-1 or TASK-3 may promote the trafficking of the channels to the surface membrane.
Rajan
Sindhu
S
Institute of Physiology, Marburg University, Deutschhausstrasse 2, 35037 Marburg, Germany.
Preisig-Müller
Regina
R
Wischmeyer
Erhard
E
Nehring
Ralf
R
Hanley
Peter J
PJ
Renigunta
Vijay
V
Musset
Boris
B
Schlichthörl
Günter
G
Derst
Christian
C
Karschin
Andreas
A
Daut
Jürgen
J
D048948:14-3-3 Proteins; D020816:Amino Acid Motifs / Q000502:physiology; D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D001692:Biological Transport / Q000502:physiology; D019556:COS Cells; D002460:Cell Line / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism; D004553:Electric Conductivity; D005260:Female; D017353:Gene Deletion; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation / Q000502:physiology; D009419:Nerve Tissue Proteins / Q000502:physiology*; D009865:Oocytes / Q000502:physiology; D010446:Peptide Fragments / Q000235:genetics; D015221:Potassium Channels / Q000502:physiology*; D024683:Potassium Channels, Tandem Pore Domain; D020033:Protein Isoforms / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D051381:Rats; D014446:Tyrosine 3-Monooxygenase / Q000502:physiology*; D014981:Xenopus
D048948:14-3-3 Proteins; C498655:KCNK9 protein, human; C407082:Kcnk9 protein, rat; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D015221:Potassium Channels; D024683:Potassium Channels, Tandem Pore Domain; D020033:Protein Isoforms; C420132:potassium channel subfamily K member 3; D014446:Tyrosine 3-Monooxygenase