Hepatocellular Transport of Biliary Phospholipids

磷脂酰胆碱转运蛋白（PC-TP）是一种类固醇生成急性调节相关的脂质转运蛋白 （START）结构域蛋白，其在体外催化磷脂酰胆碱的膜间转移， 独家。在上一个项目期间，我们证明了PC-TP在胆汁脂质中的重要作用， 分泌、肝胆固醇稳态和高密度脂蛋白（HDL）代谢。我们还解决了 复合磷脂酰胆碱的蛋白质的三维结构，并确定膜 交互作用域与我们最初预测的PC-TP作用于穿梭磷脂酰胆碱相反， 从内质网到分泌到胆汁的小管质膜，这项研究 表明在肝脏脂质稳态中具有更全面的调节作用。目前的提案测试了 假设PC-TP感知膜磷脂酰胆碱的脂肪酰基链组成， 参与控制肝脏内脂质代谢的蛋白质-蛋白质相互作用。初步 研究已经证明甘油三酯在Pctp+小鼠的肝脏中积累，并且PC-TP在小鼠的肝脏中高度蓄积。 由过氧化物酶体增殖物激活受体α（PPARa）调节。具体目标1将使用Pctp“0”~， 野生型同窝对照小鼠，以检查PC-TP对肝脏甘油三酯代谢的影响，非常 低密度脂蛋白（VLDL）的产生和调节脂质的肝脏基因的表达 体内平衡将用富含饱和脂肪的饮食激发小鼠，饱和脂肪可增加肝脏 甘油三酸酯的生产和出口，以及补充PPARct配体非诺贝特的饮食， 促进肝脏对脂肪酸的利用。具体目标2将描述PC-TP之间的相互作用 以及通过酵母双杂交筛选在小鼠肝脏和胚胎中鉴定的蛋白质。第3章将 阐明结合磷脂酰胆碱所需的PC-TP的构象变化，以及 PC-TP与其他蛋白质相互作用的结构基础。这些研究将提供 对START结构域蛋白的生物学和肝细胞凋亡的分子调控的基本新见解 脂质代谢，这可能会导致新的战略，诊断和管理， 常见疾病，如非酒精性脂肪肝。

Phosphatidylcholine transfer protein (PC-TP) is a steroidogenic acute regulatory-related lipid transfer (START) domain protein, which in vitro catalyzes the intermembrane transfer of phosphatidylcholines, exclusively. During the previous project period, we demonstrated important roles for PC-TP in biliary lipid secretion, hepatic cholesterol homeostasis and high density lipoprotein (HDL) metabolism. We also solved the three-dimensional structure of the protein complexing a phosphatidylcholine and identified the membrane interaction domain. In contrast to our original prediction that PC-TP acts to shuttle phosphatidylcholines from the endoplasmic reticulum to the canalicular plasma membrane for secretion into bile, this research has suggested a more global regulatory role in hepatic lipid homeostasis. The current proposal tests the hypothesis that PC-TP senses the fatty acyl chain composition of membrane phosphatidylcholines and engages in protein-protein interactions that control the metabolism of lipids within the liver. Preliminary studies have demonstrated that triglycerides accumulate in livers of Pctp~'~ mice and that PC-TP is highly regulated by peroxisomal proliferator-activated receptor alpha (PPARa). Specific Aim 1 will use Pctp~'~ and wild type littermate control mice to examine the influence of PC-TP on hepatic triglyceride metabolism, very low density lipoprotein (VLDL) production and the expression of hepatic genes that regulate lipid homeostasis. Mice will be challenged with a diet enriched in saturated fat,which increases hepatic production and export of triglycerides, and a diet supplemented with the PPARct ligand fenofibrate, which promotes fatty acid utilization by the liver. Specific Aim 2 will characterize the interactions between PC-TP and proteins identified in mouse liver and embryo by yeast two-hybrid screening. Specific Aim 3 will elucidate the conformational changes of PC-TP that are required to bind phosphatidylcholines, as well as the structural basis for interactions between PC-TP and other proteins. These studies should provide fundamental new insights into the biology of START domain proteins and the molecular regulation of hepatic lipid metabolism, which may potentially lead to novel strategies for the diagnosis and management of common disorders, such as nonalcoholic fatty liver disease.