Calmodulin activation limits the rate of KCNQ2 K+ channel exit from the endoplasmic reticulum.
The potential regulation of protein trafficking by calmodulin (CaM) is a novel concept that remains to be substantiated. We proposed that KCNQ2 K+ channel trafficking is regulated by CaM binding to the C-terminal A and B helices. Here we show that the L339R mutation in helix A, which is linked to human benign neonatal convulsions, perturbs CaM binding to KCNQ2 channels and prevents their correct trafficking to the plasma membrane. We used glutathione S-transferase fused to helices A and B to examine the impact of this and other mutations in helix A (I340A, I340E, A343D, and R353G) on the interaction with CaM. The process appears to require at least two steps; the first involves the transient association of CaM with KCNQ2, and in the second, the complex adopts an "active" conformation that is more stable and is that which confers the capacity to exit the endoplasmic reticulum. Significantly, the mutations that we have analyzed mainly affect the stability of the active configuration of the complex, whereas Ca2+ alone appears to affect the initial binding step. The spectrum of responses from this collection of mutants revealed a strong correlation between adopting the active conformation and channel trafficking in mammalian cells. These data are entirely consistent with the concept that CaM bound to KCNQ2 acts as a Ca2+ sensor, conferring Ca2+ dependence to the trafficking of the channel to the plasma membrane and fully explaining the requirement of CaM binding for KCNQ2 function.
Alaimo
Alessandro
A
Unidad de Biofísica, Consejo Superior de Investigaciones Cientificas-Universidad del Pais Vasco/Euskal Herriko Unibersitatea (CSIC-UPV/EHU), Spain.
Gómez-Posada
Juan Camilo
JC
Aivar
Paloma
P
Etxeberría
Ainhoa
A
Rodriguez-Alfaro
Jose Angel
JA
Areso
Pilar
P
Villarroel
Alvaro
A
D000818:Animals; D001665:Binding Sites; D002147:Calmodulin / Q000378:metabolism*; D002460:Cell Line; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006801:Humans; D051658:KCNQ2 Potassium Channel / Q000737:chemistry / Q000378:metabolism*; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D021381:Protein Transport; D051381:Rats; D013050:Spectrometry, Fluorescence; D014981:Xenopus
D002147:Calmodulin; D051658:KCNQ2 Potassium Channel; C496914:KCNQ2 protein, human; D050505:Mutant Proteins; D021122:Protein Subunits