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(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，包括参与脂肪合成的生物合成酶和参与脂肪周转的磷脂酶。脂质平衡缺陷可能导致许多疾病的发展，包括肥胖、糖尿病和癌症。在这里给出的初步结果中，我们已经确定Ypk1，人血清和糖皮质激素诱导的激酶(SGK1)的酵母同系物，是一种新的磷脂酰胆碱(PC)质膜(PM)转换调节因子。重要的是，人类SGK1修复了与ypk1突变相关的生长缺陷，表明人类和酵母蛋白之间的功能保守。PYK1酵母突变体在PM显示Plb1介导的PC脱酰化增强此外，发生在ypk1突变体中的Plb1活性加速是细胞存活的一种补偿机制，因为与ypk1突变体相比，ypk1plb1双重突变体表现出更多的生长缺陷和对细胞壁干扰剂Calcoflor的敏感性。由于PC是一种丰富的磷脂，其新陈代谢的变化可能会影响膜的组成和物理性质，从而影响其中的膜蛋白。事实上，我们已经鉴定了两种PM蛋白，它们的活性因Ypk1或Plb1的丢失而受到影响。我们的中心假设是YPK1或PLB1的缺失改变了PM的脂质组成，进而影响了PM整体蛋白的活性，导致了生长缺陷。在这里提出的研究中，我们将描述Ypk1和Plb1调节PM蛋白整体活性的过程，包括研究Plb1和Ypk1在控制PM脂组成中所起的作用。此外，对Ypk1或Plb1丢失后活性降低的PM渗透酶的脂质需求将使用单层脂泡进行检测。