Protein-Membrane Interactions in Regulated Exocytosis

调节胞吐作用中的蛋白质-膜相互作用

基本信息

批准号：
9212817
负责人：
Jingshi Shen
金额：
$ 28.98万
依托单位：
UNIVERSITY OF COLORADO
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-04-01 至 2018-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9212817
关键词：
Address Adipose tissue Binding Biochemical Biological Assay Biological Models Blood Glucose C2 Domain Cell membrane Cell surface Cells Chimeric Proteins Complex Data Development Diabetes Mellitus Disease Electrons Elements Epilepsy Event Exocytosis GLUT4 gene Glucose Transporter Goals Human Hydrophobicity Immune System Diseases Individual Insulin Intracellular Membranes Kinetics Knowledge Lipid Bilayers Lipids Mediating Membrane Membrane Fusion Membrane Proteins Molecular Molecular Mechanisms of Action Munc18c protein Muscle Fibers Non-Insulin-Dependent Diabetes Mellitus Pathway interactions Phosphorylation Physiological Physiological Processes Physiology Process Proteins Pulmonary Surfactants Reaction Research Role S-nitro-N-acetylpenicillamine SNAP receptor Site Stimulus Stress System T-Lymphocyte Therapeutic Intervention Vesicle Work base biophysical properties blood glucose regulation human disease in vivo insight microscopic imaging mutant novel novel therapeutic intervention novel therapeutics proteoliposomes public health relevance receptor reconstitution syntaxin 4 syntaxin binding protein 1 trafficking uptake vesicular SNARE proteins

项目摘要

DESCRIPTION (provided by applicant): Regulated exocytosis is a stimulus-dependent membrane fusion event of fundamental importance to a range of physiological processes. The membrane fusion reaction involves substantial lipid rearrangements and is opposed by the powerful hydrophobic force. For fusion to occur, a high energy barrier must be overcome. The overall goal of this proposed research is to determine the functional roles of membrane bilayer remodeling in overcoming the energy barrier of exocytic vesicle fusion, using the trafficking of the glucose transporter GLUT4 as a model system. To achieve this goal, we will employ a unique combination of complementary approaches including biochemical reconstitution, electron microscopic imaging, and biophysical measurements. First, we will define how two membrane bilayers are brought into close apposition by the SNARE-SM complex to prepare for the subsequent steps of membrane remodeling and merging. Next, we will assess the functional role of vesicle membrane bending in the fusion reaction. Finally, we will examine how the fusion reaction is regulated by local remodeling of the plasma membrane. We hypothesize that the fusion of exocytic vesicles with the plasma membrane involves a novel dual-curvature-induction mechanism: while the amphipathic motif of the v-SNARE bends the vesicle membrane, the hydrophobic loops of C2-domain molecules penetrate into the plasma membrane and induce local membrane curvature. Together, these membrane-bending activities are expected to create curvature stresses at the fusion sites to overcome the energy barrier for the fusion reaction. Successful completion of this proposed research will provide key insights into the molecular mechanisms of exocytic vesicle fusion. This work will also serve as a paradigm for understanding the general principles of intracellular membrane fusion. Ultimately, our findings will facilitate the development of novel therapeutic strategies for diseases associated with dysfunctional regulated exocytosis including diabetes, epilepsy, and immune disorders.

描述（由申请人提供）：受调节的胞吐作用是刺激依赖性的膜融合事件，对一系列生理过程的重要性。膜融合反应涉及大量的脂质重排，并与强大的疏水力相反。为了融合，必须克服高能屏障。这项拟议的研究的总体目标是使用葡萄糖转运蛋白glut4作为模型系统的运输，确定膜双层重塑在克服外囊泡融合的能屏障中的功能作用。为了实现这一目标，我们将采用独特的互补方法组合，包括生化重建，电子显微镜成像和生物物理测量。首先，我们将定义如何通过SNARE-SM Complex将两个膜双层构成近距离，以准备随后的膜重塑和合并步骤。接下来，我们将评估囊泡膜弯曲在融合反应中的功能作用。最后，我们将研究如何通过质膜的局部重塑来调节融合反应。我们假设外生囊泡与质膜的融合涉及一种新型的双膜诱导机制：V-snare的两亲性基序弯曲囊泡膜，囊泡膜，C2-核分子的疏水lo液渗透到浆膜膜上和层状层状层状层状。总之，这些膜弯曲活性有望在融合位点产生曲率应力，以克服融合反应的能屏障。这项拟议的研究的成功完成将为胞外囊泡融合的分子机制提供关键的见解。这项工作还将成为理解细胞内膜融合的一般原理的范式。最终，我们的发现将有助于发展与功能失调的调节胞吐作用有关的疾病的新型治疗策略，包括糖尿病，癫痫和免疫疾病。