Maxik Interaction with Gaba Transporter 3 and Heat Shock Protein 60 in the Mouse Brain

Large conductance voltage- and calcium-activated potassium (MaxiK) channels regulate Ca2+ signaling, transmitter release, and repolarization of the action potential in neurons. Analysis of MaxiK interactome from the mouse brain using liquid chromatography/mass spectrometry in tandem and knockout animals, as negative controls, revealed that MaxiK interacts with a variety of proteins localized to the mitochondria, plasma membrane, cytoplasm, Golgi apparatus, endoplasmic reticulum and other intracellular organelles. We examined whether proteins identified by mass spectrometry colocalize with MaxiK channel in cells. To this end, we have first chosen two proteins, namely gamma-aminobutyric acid (GABA) transporter (GAT)-3, and a heat shock protein (HSP)-60. GAT-3 is expressed in unmyelinated axons and glial processes, whereas HSP-60 is present in the cytosol and mitochondria, and is implicated in translocation of mitochondrial proteins from the cytoplasm. Immunocytochemical studies in HEK293T cells showed that MaxiK colocalizes with GAT-3 at the plasma membrane and HSP60 at the cell periphery. These results indicate that MaxiK could be playing a role in modulating GABA release from the presynaptic nerve terminals via GAT-3, and HSP-60 could be involved in translocating MaxiK to the mitochondria. Supporting the latter, we have confirmed the presence of MaxiK in isolated brain mitochondria using immunocytochemistry. Further studies will help to understand the role of MaxiK in modulating GAT-3 or vice versa and the role of HSP-60 in targeting MaxiK to brain mitochondria. Supported by AHA and NIH.