G protein-coupled receptor kinase 2-mediated phosphorylation of downstream regulatory element antagonist modulator regulates membrane trafficking of Kv4.2 potassium channel.
Downstream regulatory element antagonist modulator (DREAM)/potassium channel interacting protein (KChIP3) is a multifunctional protein of the neuronal calcium sensor subfamily of Ca2+-binding proteins with specific roles in different cell compartments. In the nucleus, DREAM acts as a Ca2+-dependent transcriptional repressor, and outside the nucleus DREAM interacts with Kv4 potassium channels, regulating their trafficking to the cell membrane and their gating properties. In this study we characterized the interaction of DREAM with GRK6 and GRK2, members of the G protein-coupled receptor kinase family of proteins, and their phosphorylation of DREAM. Ser-95 was identified as the site phosphorylated by GRK2. This phosphorylation did not modify the repressor activity of DREAM. Mutation of Ser-95 to aspartic acid, however, blocked DREAM-mediated membrane expression of the Kv4.2 potassium channel without affecting channel tetramerization. Treatment with the calcineurin inhibitors FK506 and cyclosporin A also blocked DREAM-mediated Kv4.2 channel trafficking and calcineurin de-phosphorylated GRK2-phosphorylated DREAM in vitro. Our results indicate that these two Ca2+-dependent posttranslational events regulate the activity of DREAM on Kv4.2 channel function.
Ruiz-Gomez
Ana
A
Departamento de Biología Molecular and Centro de Biología Molecular Severo Ochoa Universidad Autonoma de Madrid-Consejo Superior de Investigaciones Científicas, Campus de Cantoblanco, 28049 Madrid, Spain.
Mellström
Britt
B
Tornero
Daniel
D
Morato
Esperanza
E
Savignac
Magali
M
Holguín
Helena
H
Aurrekoetxea
Koldo
K
González
Paz
P
González-García
Carmen
C
Ceña
Valentín
V
Mayor
Federico
F
Naranjo
Jose R
JR
D019703:Calcineurin / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D054769:G-Protein-Coupled Receptor Kinase 2; D054768:G-Protein-Coupled Receptor Kinases; D006801:Humans; D007668:Kidney / Q000166:cytology; D051605:Kv Channel-Interacting Proteins / Q000378:metabolism*; D007930:Leucine / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D020836:Protein Structure, Quaternary; D021381:Protein Transport / Q000502:physiology*; D012097:Repressor Proteins / Q000378:metabolism*; D051672:Shal Potassium Channels / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity; D014158:Transcription, Genetic / Q000502:physiology; D020798:Two-Hybrid System Techniques; D051552:beta-Adrenergic Receptor Kinases / Q000378:metabolism*
C496749:KCNIP3 protein, human; D051605:Kv Channel-Interacting Proteins; D012097:Repressor Proteins; D051672:Shal Potassium Channels; D017346:Protein Serine-Threonine Kinases; C496567:GRK2 protein, human; D051552:beta-Adrenergic Receptor Kinases; D054769:G-Protein-Coupled Receptor Kinase 2; D054768:G-Protein-Coupled Receptor Kinases; C089176:G-protein-coupled receptor kinase 6; D019703:Calcineurin; D007930:Leucine; D002118:Calcium