Regulation of CETP by lipid transfer inhibitor protein

脂质转移抑制蛋白对 CETP 的调节

• 批准号: 7056766
• 负责人: RICHARD E MORTON
• 金额: $ 33.62万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-25 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7056766
• 关键词: apoptosis; blood lipoprotein biosynthesis; blood lipoprotein metabolism; cholesterol esters; clinical research; gene deletion mutation; high density lipoproteins; homeostasis; human tissue; immunoregulation; inhibitor /antagonist; laboratory rabbit; lipid biosynthesis; lipid metabolism; lipid transport; low density lipoprotein; protein binding; protein degradation; protein structure function; recombinant proteins; tissue /cell culture; transport proteins; very low density lipoprotein

'DESCRIPTION (provided by applicant): Plasma cholesterol and low density lipoprotein (LDL) concentrations are major risk factors for atherosclerosis, whereas elevated high density lipoprotein (HDL) levels decreased risk. Cholesteryl ester transfer protein (CETP) transports lipids between lipoproteins and, consequently, directly affects lipoprotein metabolism and alters LDL and HDL concentrations. CETP levels are highly responsive to dietary lipids and genetic factors. Additionally, CETP activity is regulated by lipid transfer inhibitor protein (LTIP). We have demonstrated that LTIP does not lower CETP activity generically but selectively suppresses CETP activity on LDL, and to a lesser extent on HDL2, but stimulates CETP activity on HDL3. Consequently, cholesterol flux through HDL is enhanced and plasma cholesteryl ester synthesis rates are increased. Thus LTIP tailors CETP-mediated lipid transfer events resulting in a lipoprotein profile that is different from that achieved by raising or lowering CETP levels alone. In this continuation application we will more fully define the functions of LTIP, and characterize its regulation and mechanism of action. We hypothesize that LTIP alters CETP activities to generate a more beneficial or atheroprotective lipoprotein profile. Three specific aims will be addressed: AIM 1) Define the mechanism by which LTIP inhibits CETP and identify the structural features of LTIP important for this activity. We will quantify binding kinetics of LTIP to LDL, HDL2 and HDL3 and determine how LTIP binding relates to CETP inhibition and the displacement of CETP from the lipoprotein surface, and define how LTIP binding influences the composition and structure of the lipoprotein surface. Mutagenesis studies will define regions and specific amino acids required for LTIP function. AIM 2) Determine the role of LTIP in regulating lipoprotein metabolism. The effects of adenoviral-mediated LTIP overexpression or LTIP suppression on lipoprotein composition and on VLDL and HDL metabolism will be defined in vivo in a hamster model. The effects of altered LTIP expression on prebeta-HDL formation and the capacity of plasma to promote cholesterol efflux from cells will also be quantified. Biochemical studies with reconstituted components will define the capacity of LTIP to influence specific aspects of VLDL and HDL metabolism, and provide a mechanistic basis for interpreting in vivo observations. AIM 3) Characterize the inactive LTIP complex and define its role in regulating LTIP activity. We have observed that LTIP is controlled by sequestration into an inactive complex. We will isolate this complex and define its protein and lipid components. Further, as association with this complex is dynamic, we will determine how metabolic processes and changes in plasma lipid levels alter the distribution of LTIP between active and inactive pools. Overall, these studies will provide novel insight into the control of CETP by LTIP and add to our understanding of how intravascular lipoprotein remodeling events contribute to steady-state lipoprotein concentration and composition.

描述（由申请人提供）：血浆胆固醇和低密度脂蛋白（LDL）浓度是动脉粥样硬化的主要危险因素，而高密度脂蛋白（HDL）水平升高可降低风险。胆固醇酯转移蛋白 (CETP) 在脂蛋白之间转运脂质，因此直接影响脂蛋白代谢并改变 LDL 和 HDL 浓度。 CETP 水平对饮食脂质和遗传因素高度敏感。此外，CETP 活性受脂质转移抑制蛋白 (LTIP) 调节。我们已经证明，LTIP 一般不会降低 CETP 活性，而是选择性抑制 LDL 上的 CETP 活性，并在较小程度上抑制 HDL2 上的 CETP 活性，但会刺激 HDL3 上的 CETP 活性。因此，通过 HDL 的胆固醇流量增加，血浆胆固醇酯合成率增加。因此，LTIP 定制了 CETP 介导的脂质转移事件，从而产生与单独提高或降低 CETP 水平所实现的脂蛋白谱不同的脂蛋白谱。 在此延续应用中，我们将更全面地定义 LTIP 的功能，并表征其调节和作用机制。我们假设 LTIP 改变 CETP 活性以产生更有益或具有动脉粥样硬化保护作用的脂蛋白谱。将解决三个具体目标： AIM 1) 定义 LTIP 抑制 CETP 的机制，并确定对此活动重要的 LTIP 结构特征。我们将量化 LTIP 与 LDL、HDL2 和 HDL3 的结合动力学，确定 LTIP 结合如何与 CETP 抑制和 CETP 从脂蛋白表面的置换相关，并定义 LTIP 结合如何影响脂蛋白表面的组成和结构。诱变研究将确定 LTIP 功能所需的区域和特定氨基酸。目的2)确定LTIP在脂蛋白代谢调节中的作用。腺病毒介导的 LTIP 过表达或 LTIP 抑制对脂蛋白组成以及 VLDL 和 HDL 代谢的影响将在仓鼠模型中体内定义。 LTIP 表达改变对前β-HDL 形成的影响以及血浆促进胆固醇从细胞流出的能力也将被量化。使用重组成分进行生化研究将确定 LTIP 影响 VLDL 和 HDL 代谢特定方面的能力，并为解释体内观察结果提供机制基础。目标 3) 表征非活性 LTIP 复合物并定义其在调节 LTIP 活性中的作用。我们观察到 LTIP 是通过隔离到非活性复合物中来控制的。我们将分离该复合物并定义其蛋白质和脂质成分。此外，由于与该复合物的关联是动态的，我们将确定代谢过程和血浆脂质水平的变化如何改变活性池和非活性池之间 LTIP 的分布。 总体而言，这些研究将为 LTIP 对 CETP 的控制提供新的见解，并加深我们对血管内脂蛋白重塑事件如何影响稳态脂蛋白浓度和组成的理解。