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Calmodulin Mediates Calcium-dependent Activation of the Intermediate Conductance KCa Channel,IKCa1

1999; Elsevier BV; Volume: 274; Issue: 9 Linguagem: Inglês

10.1074/jbc.274.9.5746

1083-351X

Christopher M. Fanger, Sanjiv Ghanshani, Naomi J. Logsdon, Heiko Rauer, Katalin Kálmán, Jianming Zhou, Kathy Beckingham, K. George Chandy, Michael D. Cahalan, Jayashree Aiyar,

Neuroscience and Neuropharmacology Research

Small and intermediate conductance Ca 2+ -activated K + channels play a crucial role in hyperpolarizing the membrane potential of excitable and nonexcitable cells. These channels are exquisitely sensitive to cytoplasmic Ca 2+ , yet their protein-coding regions do not contain consensus Ca 2+ -binding motifs. We investigated the involvement of an accessory protein in the Ca 2+ -dependent gating of hIKCa1 , a human intermediate conductance channel expressed in peripheral tissues. Cal- modulin was found to interact strongly with the cytoplasmic carboxyl (C)-tail of hIKCa1 in a yeast two-hybrid system. Deletion analyses defined a requirement for the first 62 amino acids of the C-tail, and the binding of calmodulin to this region did not require Ca 2+ . The C-tail of hSKCa3, a human neuronal small conductance channel, also bound calmodulin, whereas that of a voltage-gated K + channel, mKv1.3, did not. Calmodulin co-precipitated with the channel in cell lines transfected with hIKCa1, but not with mKv1.3 -transfected lines. A mutant calmodulin, defective in Ca 2+ sensing but retaining binding to the channel, dramatically reduced current amplitudes when co-expressed with hIKCa1 in mammalian cells. Co-expression with varying amounts of wild-type and mutant calmodulin resulted in a dominant-negative suppression of current, consistent with four calmodulin molecules being associated with the channel. Taken together, our results suggest that Ca 2+ -calmodulin-induced conformational changes in all four subunits are necessary for the channel to open.