MECHANISMS OF BILE PIGMENT EXCRETION

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Using molecular, cell biologic, biochemical and immunomorphologic methods, we request to continue long range studies of mechanisms involved in the transfer of various organic molecules (i.e., bile acids, pigments, drugs, metabolites and others) from blood to bile and back. In particular, we seek to determine the structure, function and regulation of four bile canalicular transporters and, ultimately, their dysfunction in developmental, acquired and inheritable cholestasis: (a) Cloning, sequencing and purification of the ATP-dependent transporters for bile acids and nonbile acid organic anions in mutant budding and fisson yeast in order to generate probes for identifying and studying structure and regulation of the mammalian homologues (which, due to low abundance, have frustrated efforts at purification and cloning). (b) Preliminary evidence suggests that mdr2, the major multidrug resistance gene product in the canaliculus, has ATP-dependent phospholipid translocase activity and increases phospholipid release from cell plasma membranes. Therefore, we seek to determine how these processes are related with respect to canalicular phospholipid transport, and the possible role of mdr2 in the "vanishing bile duct syndrome". (c) Cloning, sequencing and study of the regulation of the canalicular Na+ dependent purine transporter which conserves nucleosides in hepatocytes.