Intersubunit Interactions Control Kir Channel Inactivation

Abstract

Inward rectifier potassium (Kir) channels are expressed in numerous mammalian tissues including the pancreas, brain, heart, and skeletal muscle and play a critical role in controlling cellular excitability. Pancreatic ATP-sensitive Kir (KATP) channels are key regulators of insulin secretion as they link cellular metabolism with membrane excitability. Loss-of-function (LOF) mutations in KATP can cause human hyperinsulinism as a result of diminished activity. Several of these LOF mutations disrupt salt-bridge interactions that are located within the intracellular subunit-subunit interfaces, and result in channel activity showing a fast inactivation following ATP removal as measured with the inside-out patch clamp technique. Inactivation can be subsequently abolished by application of PIP2 to the cytoplasmic face of the membrane, an action that can be explained by a simple model in which PIP2 competes with the closed inactivated state. We find that homologous mutations in the strong inward rectifying Kir2.1 channel cause lower basal activity as a result of reduced apparent PIP2 sensitivity, indicating increased inactivation. Kir2.1 channels contain additional intersubunit salt-bridge interactions that are not present in KATP channels. Introduction of these salt-bridges into the inactivating KATP channel mutants partially rescues the channel from this inactivating phenotype. These results lead us to propose that the stability of the intersubunit interface regulates channel inactivation, sensitivity to PIP2, and is conserved across the Kir channel family and stabilizing the intersubunit interface provides a potential strategy to exploit in development of activating modulators of KATP and other Kir channels