RT Journal Article SR Electronic T1 PKA Reduces the Rat and Human KCa3.1 Current, CaM Binding, and Ca2+ Signaling, Which Requires Ser332/334 in the CaM-Binding C Terminus JF The Journal of Neuroscience JO J. Neurosci. FD Society for Neuroscience SP 13371 OP 13383 DO 10.1523/JNEUROSCI.1008-14.2014 VO 34 IS 40 A1 Raymond Wong A1 Lyanne C. Schlichter YR 2014 UL http://www.jneurosci.org/content/34/40/13371.abstract AB The Ca2+-dependent K+ channel, KCa3.1 (KCNN4/IK/SK4), is widely expressed and contributes to cell functions that include volume regulation, migration, membrane potential, and excitability. KCa3.1 is now considered a therapeutic target for several diseases, including CNS disorders involving microglial activation; thus, we need to understand how KCa3.1 function is regulated. KCa3.1 gating and trafficking require calmodulin binding to the two ends of the CaM-binding domain (CaMBD), which also contains three conserved sites for Ser/Thr kinases. Although cAMP protein kinase (PKA) signaling is important in many cells that use KCa3.1, reports of channel regulation by PKA are inconsistent. We first compared regulation by PKA of native rat KCa3.1 channels in microglia (and the microglia cell line, MLS-9) with human KCa3.1 expressed in HEK293 cells. In all three cells, PKA activation with Sp-8-Br-cAMPS decreased the current, and this was prevented by the PKA inhibitor, PKI14–22. Inhibiting PKA with Rp-8-Br-cAMPS increased the current in microglia. Mutating the single PKA site (S334A) in human KCa3.1 abolished the PKA-dependent regulation. CaM-affinity chromatography showed that CaM binding to KCa3.1 was decreased by PKA-dependent phosphorylation of S334, and this regulation was absent in the S334A mutant. Single-channel analysis showed that PKA decreased the open probability in wild-type but not S334A mutant channels. The same decrease in current for native and wild-type expressed KCa3.1 channels (but not S334A) occurred when PKA was activated through the adenosine A2a receptor. Finally, by decreasing the KCa3.1 current, PKA activation reduced Ca2+-release-activated Ca2+ entry following activation of metabotropic purinergic receptors in microglia.