Ordered Membrane Domain Formation and Function in Pathogenic Bacteria

病原菌中有序膜结构域的形成和功能

• 批准号: 8449208
• 负责人: Erwin London
• 金额: $ 28.76万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8449208
• 关键词: 7-dehydrocholesterol; Affinity; Antibodies; Bacteria; Bacterial Infections; Biochemical; Biological Process; Borrelia; Borrelia burgdorferi; Cell membrane; Cell physiology; Cells; Cholesterol; Collaborations; Complement; Detection; Disease; Drug Targeting; Duodenal Ulcer; Electron Microscopy; Ergosterol; Eukaryotic Cell; Gastric ulcer; Glycolipids; Goals; Immune; In Vitro; Infection; Lead; Lipid Bilayers; Lipids; Liquid substance; Lyme Disease; Membrane; Membrane Microdomains; Membrane Proteins; Membrane Structure and Function; Modeling; Nature; Pharmaceutical Preparations; Phase; Phospholipids; Pneumonia; Predisposition; Prevention; Property; Proteins; Relapsing Fever; Role; Signal Transduction; Sphingolipids; Sterols; Testing; Unsaturated Fats; Vesicle; Viral; Work; bactericide; base; combat; lipid disorder; membrane model; pathogenic bacteria; reconstitution; research study; saturated fat

DESCRIPTION (provided by applicant): The formation, properties and biological functions of membrane microdomains rich in cholesterol is one of the most exciting and controversial topics in membrane structure and function. In 1994 we proposed (with Deborah Brown, Stony Brook U.) the current working model for the nature of such domains in eukaryotic cells: that they are cholesterol and saturated lipid-rich liquid ordered state lipid domains that segregate from disordered lipid regions rich in unsaturated lipids. A number of pathogenic bacteria, including those in the genus Borrelia, obtain cholesterol from their hosts. In collaboration with Dr. Jorge Benach (Stony Brook U.) the co-discoverer of Borrelia burgdorferi as the causative agent of Lyme disease, we found that B. burgdorferi forms cholesterol glycolipid-containing outer membrane microdomains large enough to directly visualize by electron microscopy. The goal of this project is, in collaboration with the Benach and Brown labs, to understand the physical basis of domain organization of cholesteryl glycolipids and free cholesterol in Borrelia membranes, and to define the functional importance of these domains. Such studies will contribute greatly to our understanding of the general underlying principles of membrane domain organization and function. Electron microscopy, spectroscopic and biochemical approaches will be used to test the hypothesis that B. burgdorferi domains are self-organized ordered lipid domains. To do this, lipid substitution experiments will be used to modify B. burgdorferi membranes in order to determine whether the ability of sterols and other lipids to form ordered domains in vitro is necessary and sufficient for them to form membrane domains in B.burgdorferi cells. Next, whether only those molecules previously shown to strongly associate with ordered membrane domains in model membranes and eukaryotic cells localize within B.burgdorferi membrane domains will be determined. Then, whether B. burgdorferi outer membrane proteins influence domain formation will be determined. To test the hypothesis that interactions specifically involving cholesteryl glycolipids are crucial for domain formation, we will determine which B. burgdorferi lipids have the ability to form membrane domains in model membrane vesicles in vitro. The final aim will be to investigate the function of B. burgdorferi membrane domains. Based on preliminary studies, whether membrane domains are necessary to maintain B.burgdorferi. membrane integrity and/or antibody susceptibility, will be tested. These studies may point the way towards biomedical applications, including prevention/treatment of Lyme disease, by modifying membrane domains. Along these lines, a strategy for combating B. burgdorferi infections by using sterols that are not toxic to eukaryotic cells, but which we have very recently discovered to be toxic to B.burgdorferi cells, will be tested. Ultimately, the studies proposed in this project may lead to a new understanding of the membranes of other cholesterol-containing pathogenic bacteria, and new strategies to combat the diseases that they cause.

描述（申请人提供）：富含胆固醇的膜微区的形成、性质和生物学功能是膜结构和功能领域最令人兴奋和最具争议的话题之一。 1994 年，我们（与 Deborah Brown、Stony Brook U.）提出了真核细胞中此类结构域性质的当前工作模型：它们是胆固醇和富含饱和脂质的液体有序态脂质结构域，与富含胆固醇和饱和脂质的液体有序态脂质结构域分离。不饱和脂质。许多病原菌，包括疏螺旋体属细菌，从宿主中获取胆固醇。与莱姆病病原体伯氏疏螺旋体的共同发现者 Jorge Benach 博士（石溪大学）合作，我们发现伯氏疏螺旋体形成了含有胆固醇糖脂的外膜微区，其大小足以通过电子显微镜直接观察。该项目的目标是与 Benach 和 Brown 实验室合作，了解疏螺旋体膜中胆固醇糖脂和游离胆固醇结构域组织的物理基础，并定义这些结构域的功能重要性。此类研究将极大地有助于我们理解膜域组织和功能的一般基本原理。电子显微镜、光谱和生化方法将用于检验伯氏疏螺旋体结构域是自组织有序脂质结构域的假设。为此，将使用脂质替代实验来修饰伯氏疏螺旋体膜，以确定甾醇和其他脂质在体外形成有序结构域的能力对于它们在伯氏疏螺旋体细胞中形成膜结构域是否是必要和充分的。接下来，将确定是否只有那些先前显示与模型膜和真核细胞中的有序膜域强相关的分子位于伯氏疏螺旋体膜域内。然后，将确定伯氏疏螺旋体外膜蛋白是否影响结构域形成。为了检验特定涉及胆固醇糖脂的相互作用对于结构域形成至关重要的假设，我们将确定哪些伯氏疏螺旋体脂质具有在体外模型膜囊泡中形成膜结构域的能力。最终目标是研究伯氏疏螺旋体膜域的功能。根据初步研究，膜域是否是维持伯氏疏螺旋体所必需的。将测试膜完整性和/或抗体敏感性。这些研究可能为生物医学应用指明道路，包括通过修饰膜域来预防/治疗莱姆病。沿着这些思路，将测试一种通过使用对真核细胞无毒但我们最近发现对伯氏疏螺旋体细胞有毒的甾醇来对抗伯氏疏螺旋体感染的策略。最终，该项目提出的研究可能会导致人们对其他含胆固醇病原菌的膜有新的认识，并找到对抗它们引起的疾病的新策略。