HEPATOCELLULAR TRANSPORT OF BILIARY PHOSPHOLIPIDS

During bile formation in humans, a transhepatic flux of an average of one gram of bile salt per hour stimulates the daily biliary secretion of two grams of cholesterol together with eleven grams of phospholipids, which are enriched (greater than 95 percent) in phosphatidylcholines. Phosphatidylcholine transfer protein is a cytosolic protein of unknown physiological function that is enriched in liver and in vitro catalyzes intermembrane transfer of phosphatidylcholines. Submicellar bile salts markedly enhance activity of purified phosphatidylcholine transfer protein and suggest a critical role for this protein in hepatocellular selection and transport of biliary phospholipids. Recent cDNA cloning has demonstrated phosphatidylcholine transfer protein to be novel and conserved among vertebrates. Insights into cellular function will be gleaned by examining in tissue culture the influence of phosphatidylcholine transfer protein on cellular secretion and uptake of lipid. To elucidate the molecular basis for its substrate specificity, x-ray crystallography will be utilized to solve the three dimensional structure of recombinant expressed phosphatidylcholine transfer protein. Structural data will be employed as a basis for structure-function analysis by site-directed mutagenesis. Preliminary studies suggest that cis acting elements in the phosphatidylcholine transfer protein gene promoter are responsible for liver-specific transcriptional regulation. Regulatory elements and transcription factors responsible for hepatic enrichment with phosphatidylcholine transfer protein will be elucidated in tissue culture. Genetic mapping of the phosphatidylcholine transfer protein gene to human chromosome 17q21-22 has revealed co-localization with Meckel syndrome, a lethal disorder characterized by multiple congenital anomalies including periportal hepatic fibrosis and bile duct proliferation in utero. Because preliminary experiments have revealed mutations in cDNAs encoding phosphatidylcholine transfer protein from patients with Meckel syndrome, a systematic study will be undertaken to test for genetic mutations in a series of patients. Influence of mutations on activity and structure of recombinant protein will be assessed. Studies of the cellular function, structural biology, molecular regulation and genetics of phosphatidylcholine transfer protein should help to elucidate biliary lipid secretion at a molecular level and potentially lead to early interventions in cholelithiasis and cholestasis, as well as new strategies for management of hypercholesterolemia.