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Intracellular traffic of the K+ channels TASK-1 and TASK-3: role of N- and C-terminal sorting signals and interaction with 14-3-3 proteins.
- Source :
-
The Journal of physiology [J Physiol] 2009 Mar 01; Vol. 587 (Pt 5), pp. 929-52. Date of Electronic Publication: 2009 Jan 12. - Publication Year :
- 2009
-
Abstract
- The two-pore-domain potassium channels TASK-1 (KCNK3) and TASK-3 (KCNK9) modulate the electrical activity of neurons and many other cell types. We expressed TASK-1, TASK-3 and related reporter constructs in Xenopus oocytes, mammalian cell lines and various yeast strains to study the mechanisms controlling their transport to the surface membrane and the role of 14-3-3 proteins. We measured potassium currents with the voltage-clamp technique and fused N- and C-terminal fragments of the channels to various reporter proteins to study changes in subcellular localisation and surface expression. Mutational analysis showed that binding of 14-3-3 proteins to the extreme C-terminus of TASK-1 and TASK-3 masks a tri-basic motif, KRR, which differs in several important aspects from canonical arginine-based (RxR) or lysine-based (KKxx) retention signals. Pulldown experiments with GST fusion proteins showed that the KRR motif in the C-terminus of TASK-3 channels was able to bind to COPI coatomer. Disabling the binding of 14-3-3, which exposes the KRR motif, caused localisation of the GFP-tagged channel protein mainly to the Golgi complex. TASK-1 and TASK-3 also possess a di-basic N-terminal retention signal, KR, whose function was found to be independent of the binding of 14-3-3. Suppression of channel surface expression with dominant-negative channel mutants revealed that interaction with 14-3-3 has no significant effect on the dimeric assembly of the channels. Our results give a comprehensive description of the mechanisms by which 14-3-3 proteins, together with N- and C-terminal sorting signals, control the intracellular traffic of TASK-1 and TASK-3.
- Subjects :
- 14-3-3 Proteins metabolism
Amino Acid Motifs genetics
Amino Acid Sequence
Animals
Female
Humans
Intracellular Space genetics
Molecular Sequence Data
Nerve Tissue Proteins genetics
Oocytes metabolism
Oocytes physiology
Potassium Channels, Tandem Pore Domain genetics
Protein Sorting Signals genetics
Protein Transport genetics
Xenopus laevis
14-3-3 Proteins physiology
Intracellular Space physiology
Nerve Tissue Proteins metabolism
Potassium Channels, Tandem Pore Domain metabolism
Protein Sorting Signals physiology
Subjects
Details
- Language :
- English
- ISSN :
- 1469-7793
- Volume :
- 587
- Issue :
- Pt 5
- Database :
- MEDLINE
- Journal :
- The Journal of physiology
- Publication Type :
- Academic Journal
- Accession number :
- 19139046
- Full Text :
- https://doi.org/10.1113/jphysiol.2008.164756