Hormone Regulation of [Ca2+]i in Pancreatic Acinar Cells

胰腺腺泡细胞中[Ca2]i的激素调节

• 批准号: 7484306
• 负责人: Shmuel Muallem
• 金额: $ 34.07万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-09-01 至 2009-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7484306
• 关键词: Achievement; Acinar Cell; Acinus organ component; Acute; Address; Animal Model; Apical; Apoptosis; Binding; Biochemical; Biological Models; Bombesin Receptor; Carrier Proteins; Cell Death; Cell physiology; Cells; Characteristics; Cholecystokinin; Collaborations; Communication; Complex; Data; Diabetes Mellitus; Disease; Dominant-Negative Mutation; Ensure; Enzymes; Functional disorder; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; G-substrate; GTP-Binding Proteins; Gene Expression; Generations; Genes; Genetic Polymorphism; Goals; Homer 1; Homer protein; Hormones; Human Resources; Hypertension; Invasive; Lead; Life; Lipids; Liquid substance; Maps; Mediating; Modeling; Mus; Mutant Strains Mice; Mutation; N-terminal; Nature; Neurotransmitters; Organelles; Pancreas; Pancreatitis; Peptides; Physiological; Play; Protein Binding; Protein Biosynthesis; Proteins; RGS Proteins; RNA Splicing; Rattus; Regulation; Role; Scaffolding Protein; Second Messenger Systems; Secretory Cell; Secretory Vesicles; Signal Transduction; Signaling Protein; Site; Specificity; TRP channel; Techniques; Tertiary Protein Structure; Testing; Time; Variant; Vascular Diseases; acute pancreatitis; base; chronic pancreatitis; cupidin; hormone regulation; in vivo; insight; mutant; protein function; receptor; scaffold; second messenger; spinophilin; tool

DESCRIPTION (provided by applicant): The polarized nature of pancreatic acinar cell functions require polarized organization and function of signaling complexes. Understanding assembly and regulation of signaling proteins within complexes in cellular microdomains is the central theme of this proposal. Based on the findings of polarized expression of Ca2+ signaling proteins in secretory cells, the discovery of direct regulation of IP3Rs channel activity by GBy, binding of RGS proteins and the third intracellular loop of GPCRs (31L) to the scaffolding protein spinophilin, the role of Homer proteins in mediated communication between TRPC channels and IP3Rs at the PM/ER junction and the mislocalization of Ca2+ transporting proteins and organelles in PERK1-/- and MISTI -/- mice, we will test the central hypothesis by I. Study gating of IP3Rs by G(( and its relevance to Ca 2+ signal in pancreatic acini. This new mode of Ca 2+ signaling by GPCRs will be examined by identifying the IP3Rs and G(( domains that mediate the regulation and the mechanism of their interaction. More important, the domains and peptides and mutations thereof will be used to determine the significance of this mode of Ca2+ signaling to Ca2+ signaling by pancreatic acini. 2. Study the role of spinophilin (SPL) in mediating communication between RGS proteins and GPCRs, This will be achieved by refining the SPL domains that interact with RGS proteins, the role of SPL in RGS proteins GAP activity and identify dominant negatives that interfere with the interaction of SPL with RGS proteins and GPCRs. These tools will be used to examine the role of SPL interactions in vivo. In a second approach, constitutively active and signaling impaired mutants in the 31L of the M3R will be used to further map the site of communication between RGS proteins and GPCR. 3. Study the role of Homer proteins in Ca 2+ signaling at the PM/ER junction. This will be achieved by examining the role of Homers in activation of TRPCI, TRPC3 and TRPC6 that are expressed in pancreatic acini by IP3Rs and by lipids. In another approach, Ca2+ signaling will be characterized in pancreatic acini of Homer I -/- mice. 4. Characterize Ca 2+ signaling in PERK1 -I-and MIST1-I- mice. Ca2+ transporting proteins and organelles are mislocalized in pancreatic acini of these mice, offering us the first opportunity to examine the role of assembly of Ca2+ signaling complexes in cellular microdomains in the initiation, propagation and generation of polarized Ca2+ waves. More important, the pancreas of the mutant mice develops with time morphological characteristics of pancreatitis. Hence, these mice may provide an animal model to study the role of aberrant Ca 2+ signaling in chronic pancreatitis. I believe that the tools available in my lab, the expertise of the personnel and the collaborations we established will allow us to achieve our goals and provide insights of general relevance to cell signaling and to the function and dysfunction of the pancreas.

描述（由申请人提供）：胰腺腺泡细胞功能的极化性质需要信号复合物的极化组织和功能。了解细胞微域复合物内信号蛋白的组装和调节是该提案的中心主题。基于分泌细胞中 Ca2+ 信号蛋白极化表达的发现、GBy 对 IP3R 通道活性的直接调节、RGS 蛋白和 GPCR 的第三个细胞内环 (31L) 与支架蛋白亲旋蛋白的结合的发现、 Homer 蛋白介导 PM/ER 连接处 TRPC 通道和 IP3R 之间的通讯以及 PERK1-/- 和 PERK1-/- 中 Ca2+ 转运蛋白和细胞器的错误定位MISTI -/- 小鼠，我们将测试 I 的中心假设。研究 G(( 的 IP3R 门控及其与胰腺腺泡中 Ca 2+ 信号的相关性。GPCR 的这种新的 Ca 2+ 信号传导模式将通过识别来检查介导调节及其相互作用机制的 IP3R 和 G(( 结构域。更重要的是，这些结构域和肽及其突变将用于确定这种 Ca2+ 信号传导模式对胰腺 Ca2+ 信号传导的重要性2. 研究亲旋蛋白 (SPL) 在介导 RGS 蛋白和 GPCR 之间的通讯中的作用，这将通过细化与 RGS 蛋白相互作用的 SPL 结构域、SPL 在 RGS 蛋白 GAP 活性中的作用并识别显性失活来实现。干扰 SPL 与 RGS 蛋白和 GPCR 的相互作用。这些工具将用于检查 SPL 相互作用在体内的作用。在第二种方法中，M3R 31L 中的组成型活性和信号传导受损的突变体将用于进一步绘制 RGS 蛋白和 GPCR 之间的通讯位点。 3. 研究 Homer 蛋白在 PM/ER 连接处 Ca 2+ 信号传导中的作用。这将通过检查 Homers 在 TRPCI、TRPC3 和 TRPC6 激活中的作用来实现，这些物质通过 IP3R 和脂质在胰腺腺泡中表达。在另一种方法中，将在 Homer I -/- 小鼠的胰腺腺泡中表征 Ca2+ 信号传导。 4. 表征 PERK1 -I- 和 MIST1-I- 小鼠中的 Ca 2+ 信号传导。 Ca2+ 转运蛋白和细胞器在这些小鼠的胰腺腺泡中错误定位，为我们提供了第一个机会来研究细胞微域中 Ca2+ 信号复合物的组装在极化 Ca2+ 波的启动、传播和生成中的作用。更重要的是，突变小鼠的胰腺随时间的发展呈现出胰腺炎的形态特征。因此，这些小鼠可以提供动物模型来研究异常Ca 2+ 信号传导在慢性胰腺炎中的作用。我相信，我的实验室提供的工具、人员的专业知识以及我们建立的合作将使我们能够实现我们的目标，并提供与细胞信号传导以及胰腺功能和功能障碍普遍相关的见解。