Molecular Mechanisms of Protein-Membrane Interactions Driving Insulin Secretion

蛋白质-膜相互作用促进胰岛素分泌的分子机制 项目概要/摘要： 这项研究计划将确定蛋白质-膜相互作用的分子机制 胰岛素分泌的核心，包括这些相互作用如何受到细胞内脂质变化的影响 与胰岛素分泌信号有关。膜靶向蛋白是胰岛素分泌的核心， 胰岛细胞，以及许多其他细胞类型的信号。一个关键 在分泌中起核心作用的膜靶向蛋白是突触结合蛋白（Syt），其具有两个C2 结构域，其通过对接到膜来触发分泌囊泡-质膜融合， 增加细胞内[Ca 2 +]。大多数探索Syt活性的分子机制的研究都集中在 Syt 1，主要Syt亚型，负责快速神经递质释放。然而，Syt 1并不扮演 在胰岛素分泌中起主要作用;相反，细胞使用Syt 7，Syt 9和Syt样蛋白颗粒亲蛋白来分泌胰岛素。 分泌途径本文提出的研究将检验来自Syt 7的C2结构域， Syt 9和颗粒亲蛋白具有专门用于其在胰岛素中的作用的生物化学和生物物理特性 分泌，因此尽管具有同源结构，但其行为方式与Syt 1不同。之前 结果支持这一论断，因为Syt 7的C2 A结构域以更强的结合力结合膜。 来自疏水效应的贡献大于来自Syt 1的相应结构域。拟议的研究将 使用已建立的生物化学和生物物理技术的组合，沿着尖端的单- 分子荧光显微镜，以探测驱动力和分子相互作用的基础上， 这些C2结构域的活性驱动胰岛素分泌。为了将这些机械研究与 细胞功能和疾病，C2结构域对信号脂质和氧化产物的敏感性也将 追究在葡萄糖刺激的胰岛素分泌过程中， 分泌，以及主要信号脂质对Syt C2结构域膜结合的影响将在这里测量。 适当控制C2结构域膜相互作用对胰岛素分泌至关重要，并且C2结构域膜相互作用的长期改变对胰岛素分泌至关重要。 这些机制可能导致伴随2型糖尿病的细胞分泌功能的丧失。 总的来说，这些结果将有助于更好地了解胰岛素分泌途径，并阐明 糖尿病细胞缺陷的潜在机制

Molecular Mechanisms of Protein-Membrane Interactions Driving Insulin Secretion Project Summary/Abstract: This research project will determine the molecular mechanisms underlying protein-membrane interactions central to insulin secretion, including how these interactions are influenced by intracellular lipid changes associated with insulin secretory signaling. Membrane-targeting proteins are central to insulin secretion in the ¿ cells of the pancreatic islets of Langerhans, as well as signaling in many other cell types. One key membrane-targeting protein that plays a central role in secretion is synaptotagmin (Syt), which has two C2 domains that trigger secretory vesicle-plasma membrane fusion by docking to membranes in response to increased intracellular [Ca2+]. Most studies probing the molecular mechanisms of Syt activity have focused on Syt1, the major Syt isoform responsible for rapid neurotransmitter release. However, Syt1 does not play a major role in insulin secretion; rather, ¿ cells use Syt7, Syt9, and the Syt-like protein granuphilin for the insulin secretion pathway. The research proposed herein will test the hypothesis that the C2 domains from Syt7, Syt9, and granuphilin have biochemical and biophysical properties specialized for their roles in insulin secretion, and thus behave in a manner distinct from Syt1 despite having homologous structures. Prior results support this assertion, as the C2A domain from Syt7 binds membranes with a much stronger contribution from the hydrophobic effect than the corresponding domain from Syt1. The proposed studies will use a combination of established biochemical and biophysical techniques, along with cutting-edge single- molecule fluorescence microscopy, to probe the driving forces and molecular interactions underlying the activities of these C2 domains driving insulin secretion. In order to connect these mechanistic studies to cellular function and disease, the sensitivity of C2 domains to signaling lipids and oxidation products will also be investigated. A number of lipid signaling pathways are activated during glucose-stimulated insulin secretion, and the effects of major signaling lipids on Syt C2 domain membrane binding will be measured here. Proper control of C2 domain membrane interactions is vital for insulin secretion, and long-term alterations in these mechanisms could contribute to the loss of ¿ cell secretory function that accompanies type 2 diabetes. Overall, the results will both lead to a better understanding of insulin secretion pathways and shed light on possible mechanisms underlying ¿ cell defects in diabetes.