a dual polybasic motif determines phosphoinositide binding and regulation in the p2x channel family双多元主题确定磷酸肌醇在家庭p2x通道绑定和监管.pdfVIP

A Dual Polybasic Motif Determines Phosphoinositide Binding and Regulation in the P2X Channel Family 1 1 ´ ´ 2 ´ ´ 1 Louis-Philippe Bernier , Dominique Blais , Eric Boue-Grabot , Philippe Seguela * ´ ´ 1 Department of Neurology and Neurosurgery, Montreal Neurological Institute, Alan Edwards Centre for Research on Pain, McGill University, Montreal, Quebec, Canada, ´ ´ ´ ´ 2 Institut des Maladies Neurodegeneratives, Universite Bordeaux Segalen, CNRS UMR 5293, Bordeaux, France Abstract Phosphoinositides modulate the function of several ion channels, including most ATP-gated P2X receptor channels in neurons and glia, but little is known about the underlying molecular mechanism. We identified a phosphoinositide-binding motif formed of two clusters of positively charged amino acids located on the P2X cytosolic C-terminal domain, proximal to the second transmembrane domain. For all known P2X subtypes, the specific arrangement of basic residues in these semi- conserved clusters determines their sensitivity to membrane phospholipids. Neutralization of these positive charges disrupts the functional properties of the prototypical phosphoinositide-binding P2X4 subtype, mimicking wortmannin- induced phosphoinositide depletion, whereas adding basic residues at homologous positions to the natively insensitive P2X5 subtype establishes de novo phosphoinositide-mediated regulation. Moreover, biochemical evidence of in vitro P2X subunit-phospholipid interaction and functional intracellular phosphoinositide-binding