MOLECULAR DETERMINANTS FOR ASSEMBLY OF G-PROTEIN-ACTIVATED INWARDLY RECTIFYING K+ CHANNELS

Kir3.1 and Kir3.2 associate to form G-protein-activated, inwardly rect ifying K+ channels. To identify regions involved in the coassembly of these subunits, truncated Kir3.1 polypeptides were coexpressed with ep itope-tagged subunits in an in vitro translation system, N-terminal, C -terminal, and core region polypeptides were coimmunoprecipitated with both Kir3.2 and Kir3.1, suggesting that multiple elements distributed throughout the Kir3.1 polypeptide contribute to intersubunit binding interactions. The Kir3.2 C-terminal polypeptide coimmunoprecipitated w ith the Kir3.1 C-terminal polypeptide, but neither region recognized t he N-terminal domain and core region of the Kir3.1 subunit. This sugge sts that within Kir3 channels the C-terminal domains of neighboring su bunits interact. Coexpression of the truncated polypeptides with Kir3. 1 and Kir3.2 in Xenopus oocytes reduced functional expression of the h eteromeric channels. Constructs encoding the core region plus N-termin al and proximal C-terminal regions competed more effectively than the core region alone, which supports the contribution of all three region s to intersubunit binding interactions. Proximal and distal segments o f the C-terminal domain were as effective at inhibiting functional exp ression as the entire C-terminal domain.