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Abstract:

The amino-terminal and carboxy-terminal domains of inwardly rectifying potassium channel (Kir) subunits are both intracellular. A direct physical interaction between these two domains is involved in the response of Kir channels to regulatory factors such as G-proteins, nucleotides and intracellular pH. We have previously mapped the region within the N-terminal domain of Kir6.2 that interacts with the C-terminus. In this study we use a similar in vitro protein-protein interaction assay to map the regions within the C-terminus which interact with the N-terminus. We find that multiple interaction domains exist within the C-terminus: CID1 (amino acids (aa) 279-323), CID2 (aa 214-222) and CID3 (aa 170-204). These domains correlate with regions previously identified as making important contributions to Kir channel assembly and function. The highly conserved nature of the C-terminus suggests that a similar association with the N-terminus may be a feature common to all members of the Kir family of potassium channels, and that it may be involved in gating of Kir channels by intracellular ligands.

Abstract:

The activity of voltage-gated potassium (Kv) channels can be dynamically modulated by several events, including neurotransmitter-stimulated biochemical cascades mediated by G-protein-coupled receptors. By using a heterologous expression system, we show that activating the 5-HT2C receptor inhibits both Kv1.1 and Kv1.2 channels through a tyrosine phosphorylation mechanism. The major molecular determinants of channel inhibition were identified as two tyrosine residues located in the N-terminal region of the Kv channel subunit. Furthermore, we demonstrate that receptor protein tyrosine phosphatase alpha (RPTPalpha), a receptor protein tyrosine phosphatase, co-ordinates the inhibition process mediated via 5-HT2C receptors. We therefore propose that the serotonergic regulation of human Kv1.1 and Kv1.2 channel activity by the 5-HT2C receptor involves the dual coordination of both RPTPalpha and specific tyrosine kinases coupled to this receptor.