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
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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. / Zuzarte, Marylou; Heusser, Katja; Renigunta, Vijay; Schlichthörl, Günter; Rinné, Susanne; Wischmeyer, Erhard; Daut, Jürgen; Schwappach, Blanche; Preisig-Müller, Regina.
in: J PHYSIOL-LONDON, Jahrgang 587, Nr. Pt 5, 01.03.2009, S. 929-52.Publikationen: SCORING: Beitrag in Fachzeitschrift/Zeitung › SCORING: Zeitschriftenaufsatz › Forschung › Begutachtung
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TY - JOUR
T1 - 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
AU - Zuzarte, Marylou
AU - Heusser, Katja
AU - Renigunta, Vijay
AU - Schlichthörl, Günter
AU - Rinné, Susanne
AU - Wischmeyer, Erhard
AU - Daut, Jürgen
AU - Schwappach, Blanche
AU - Preisig-Müller, Regina
PY - 2009/3/1
Y1 - 2009/3/1
N2 - 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.
AB - 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.
KW - 14-3-3 Proteins/metabolism
KW - Amino Acid Motifs/genetics
KW - Amino Acid Sequence
KW - Animals
KW - Female
KW - Humans
KW - Intracellular Space/genetics
KW - Molecular Sequence Data
KW - Nerve Tissue Proteins/genetics
KW - Oocytes/metabolism
KW - Potassium Channels, Tandem Pore Domain/genetics
KW - Protein Sorting Signals/genetics
KW - Protein Transport/genetics
KW - Xenopus laevis
U2 - 10.1113/jphysiol.2008.164756
DO - 10.1113/jphysiol.2008.164756
M3 - SCORING: Journal article
C2 - 19139046
VL - 587
SP - 929
EP - 952
JO - J PHYSIOL-LONDON
JF - J PHYSIOL-LONDON
SN - 0022-3751
IS - Pt 5
ER -