Potassium channel silencing by constitutive endocytosis and intracellular sequestration.
Tandem of P domains in a weak inwardly rectifying K(+) channel 1 (TWIK1) is a K(+) channel that produces unusually low levels of current. Replacement of lysine 274 by a glutamic acid (K274E) is associated with stronger currents. This mutation would prevent conjugation of a small ubiquitin modifier peptide to Lys-274, a mechanism proposed to be responsible for channel silencing. However, we found no biochemical evidence of TWIK1 sumoylation, and we showed that the conservative change K274R did not increase current, suggesting that K274E modifies TWIK1 gating through a charge effect. Now we rule out an eventual effect of K274E on TWIK1 trafficking, and we provide convincing evidence that TWIK1 silencing results from its rapid retrieval from the cell surface. TWIK1 is internalized via a dynamin-dependent mechanism and addressed to the recycling endosomal compartment. Mutation of a diisoleucine repeat located in its cytoplasmic C terminus (I293A,I294A) stabilizes TWIK1 at the plasma membrane, resulting in robust currents. The effects of I293A,I294A on channel trafficking and of K274E on channel activity are cumulative, promoting even more currents. Activation of serotoninergic receptor 5-HT(1)R or adrenoreceptor alpha2A-AR stimulates TWIK1 but has no effect on TWIK1I293A,I294A, suggesting that G(i) protein activation is a physiological signal for increasing the number of active channels at the plasma membrane.
Feliciangeli
Sylvain
S
Institut de Pharmacologie Moléculaire et Cellulaire, CNRS, and Université de Nice Sophia-Antipolis, Sophia-Antipolis, 06560 Valbonne, France.
Tardy
Magalie P
MP
Sandoz
Guillaume
G
Chatelain
Franck C
FC
Warth
Richard
R
Barhanin
Jacques
J
Bendahhou
Saïd
S
Lesage
Florian
F
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004285:Dogs; D004594:Electrophysiology; D004705:Endocytosis / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007150:Immunohistochemistry; D008854:Microscopy, Electron; D009154:Mutation; D010766:Phosphorylation / Q000187:drug effects; D024683:Potassium Channels, Tandem Pore Domain / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000235:genetics / Q000502:physiology*; D044263:Receptors, Serotonin, 5-HT1 / Q000378:metabolism; D012701:Serotonin / Q000494:pharmacology
C109137:KCNK1 protein, human; D024683:Potassium Channels, Tandem Pore Domain; D044263:Receptors, Serotonin, 5-HT1; D012701:Serotonin