Inhibition of BK channel activity by association with calcineurin in rat brain.
Large conductance calcium-activated potassium (BK(Ca)) channels are regulated by a number of different protein kinases and phosphatases. The close association of enzymes and channel have been shown to underlie many examples of modulation. However, only the association of protein kinase A with the BK(Ca) channel has been detailed [Tian et al. (2003)J. Biol. Chem., 278, 8669-8677]. We have found using reciprocal immunoprecipitations that the BK(Ca) channel associates with the calcium/calmodulin-dependent phosphatase calcineurin, in Wistar rat brain. A HA-tagged construct of the carboxyl terminus of rSlo(27), a variant of the BK(Ca) channel that is abundant in the hippocampus [Ha et al. (2000)Eur. J. Biochem., 267, 910-9218], was found to associate only with the B subunit of calcineurin. This data suggests that the majority of the interaction of the BK(Ca) channel with calcineurin is mediated by the B subunit of the phosphatase. This was confirmed by using glutathione-S-transferase (GST) fusion proteins of the linker regions between the S7-S10 hydrophobic domains in the carboxyl terminus of rSlo(27), where only the B subunit of calcineurin interacted with regions between S7 and S9 of the channel. Addition of a constitutively active calcineurin (CaN(420)) to inside-out membrane patches excised from cultured hippocampal neurons resulted in a dramatic reduction in BK(Ca) channel open probability, with only very short-duration events being apparent. These data suggest that BK(Ca) channel activity is inhibited by calcineurin, an effect mediated by the association of the calcineurin B subunit with the carboxyl terminus of the channel.
Loane
David J
DJ
Department of Pharmacology and MRC Centre for Synaptic Plasticity, School of Medical Sciences, University of Bristol, Bristol, BS8 1TD, UK.
Hicks
Gareth A
GA
Perrino
Brian A
BA
Marrion
Neil V
NV
D000818:Animals; D000831:Animals, Newborn; D001921:Brain / Q000378:metabolism*; D019703:Calcineurin / Q000737:chemistry / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009474:Neurons / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D017434:Protein Structure, Tertiary / Q000187:drug effects / Q000502:physiology; D021122:Protein Subunits / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D051036:Large-Conductance Calcium-Activated Potassium Channels; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D019703:Calcineurin