The Role of Phospholipid Turnover in Membrane Protein Function

磷脂周转在膜蛋白功能中的作用

• 批准号: 8434430
• 负责人: JANA L VOGT
• 金额: $ 30.36万
• 依托单位: DUQUESNE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-12-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8434430
• 关键词: Address; Affect; Biochemical; Cell Survival; Cell Wall; Cell membrane; Cell physiology; Cells; Defect; Development; Diabetes Mellitus; Disease; Environment; Enzymes; Exhibits; Glucans; Glucocorticoids; Growth; Homeostasis; Homologous Gene; Human; In Vitro; Lecithin; Lipids; Liposomes; Lysophospholipase; Malignant Neoplasms; Mammalian Cell; Mediating; Membrane; Membrane Lipids; Membrane Proteins; Metabolic; Metabolic syndrome; Metabolism; Microscopic; Modeling; Molecular Genetics; Obesity; Performance; Phospholipase; Phospholipids; Phosphotransferases; Play; Process; Proteins; Research Personnel; Role; Serum; Sphingolipids; System; Techniques; Testing; Vesicle; Yeasts; deacylation; enzyme activity; glucan synthase; glycerylphosphoinositol; graduate student; in vitro activity; in vivo; interest; mutant; novel; overexpression; permease; physical property; protein function; proteoliposomes; public health relevance; reconstitution; research study; response; undergraduate student; yeast protein

DESCRIPTION (provided by applicant): Membrane homeostasis is a fundamental aspect of cell function and requires the coordinated control of multiple metabolic activities, including biosynthetic enzymes involved in lipid synthesis and phospholipases involved in lipid turnover. Defects in lipid homeostasis can contribute to the development of many diseases, including obesity, diabetes, and cancer. In preliminary results presented here, we have identified Ypk1, the yeast homolog of the human serum and glucocorticoid-induced kinase (SGK1), as a novel regulator of phosphatidylcholine (PC) turnover at the plasma membrane (PM). Importantly, human SGK1 rescues growth defects associated with a ypk1 mutant, indicating functional conservation between the human and yeast proteins. A ypk1 yeast mutant exhibits increased Plb1-mediated PC deacylation at the PM. Furthermore, the accelerated Plb1 activity that occurs in a ypk1 mutant is a compensatory mechanism for cell survival, as a ypk1plb1 double mutant exhibits an increased growth defect and increased sensitivity to the cell wall perturbing agent, calcofluor, as compared to the ypk1 mutant. Because PC is an abundant phospholipid, alterations in its metabolism might be expected to affect the composition and physical properties of the membrane, and, therefore, membrane proteins that reside within. Indeed, we have identified two PM proteins whose activities are compromised by loss of Ypk1 or Plb1. Our central hypothesis is that deletion of YPK1 or PLB1 alters PM lipid composition, which, in turn, impinges upon the activity of integral PM proteins and results in growth defects. In the studies proposed here, we will delineate the process by which Ypk1 and Plb1 modulate integral PM protein activity, including examining the role that Plb1 and Ypk1 play in controlling PM lipid composition. In addition, the lipid requirements of a PM permease whose activity is reduced in response to loss of Ypk1 or Plb1 will be examined using unilamellar lipid vesicles.

描述（由申请人提供）：膜稳态是细胞功能的一个基本方面，需要协调控制多种代谢活动，包括参与脂质合成的生物合成酶和参与脂质周转的磷脂酶。脂质稳态缺陷可能导致许多疾病的发生，包括肥胖、糖尿病和癌症。在此介绍的初步结果中，我们已确定 Ypk1（人血清和糖皮质激素诱导激酶 (SGK1) 的酵母同源物）作为质膜 (PM) 磷脂酰胆碱 (PC) 周转的新型调节剂。重要的是，人类 SGK1 可以挽救与 ypk1 突变体相关的生长缺陷，这表明人类和酵母蛋白之间存在功能保守性。 ypk1 酵母突变体在 PM 处表现出 Plb1 介导的 PC 脱酰作用增加。此外，ypk1 突变体中 Plb1 活性的加速是细胞存活的一种补偿机制，因为与 ypk1 突变体相比，ypk1plb1 双突变体表现出生长缺陷增加和对细胞壁扰动剂 calcoflor 的敏感性增加。由于 PC 是一种丰富的磷脂，其代谢的改变可能会影响膜的组成和物理特性，从而影响膜内的膜蛋白。事实上，我们已经鉴定出两种 PM 蛋白，它们的活性因 Ypk1 或 Plb1 的丢失而受到损害。我们的中心假设是，YPK1 或 PLB1 的缺失会改变 PM 脂质成分，进而影响完整 PM 蛋白的活性并导致生长缺陷。在此提出的研究中，我们将描述 Ypk1 和 Plb1 调节整体 PM 蛋白活性的过程，包括检查 Plb1 和 Ypk1 在控制 PM 脂质组成中所起的作用。此外，将使用单层脂质囊泡检查 PM 通透酶的脂质需求，该酶的活性因 Ypk1 或 Plb1 的丢失而降低。