Preproinsulin Translocation and Processing at the ER Membrane: a link to Diabetes

内质网膜上的前胰岛素原易位和加工：与糖尿病的联系

• 批准号: 8842623
• 负责人: Ming Liu
• 金额: $ 27.21万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8842623
• 关键词: Affect; Alleles; Alzheimer' s Disease; American; Anabolism; Automobile Driving; Behavior; Beta Cell; Cells; Clinical; Code; Computational Biology; Data; Defect; Development; Diabetes Mellitus; Dimerization; Disease; Docking; Dominant-Negative Mutation; Elements; Endoplasmic Reticulum; Event; Exposure to; Failure; Functional disorder; Genes; Goals; Heterozygote; Human; Insulin; Insulin Resistance; Investigation; Investigational Therapies; Kinetics; Lead; Life; Link; Membrane; Modeling; Molecular; Mutant Strains Mice; Mutation; Neonatal; Neurodegenerative Disorders; Non-Insulin-Dependent Diabetes Mellitus; Pancreas; Pathogenesis; Patients; Peptide Signal Sequences; Phenotype; Play; Population; Process; Production; Proinsulin; Relative (related person); Reporting; Role; Secretory Vesicles; Series; Site; Structure; Structure of beta Cell of islet; Testing; Time; Toxic effect; Validation; base; design; endoplasmic reticulum stress; gain of function; genetic manipulation; intermolecular interaction; mutant; neonatal diabetes mellitus; novel; novel strategies; preproinsulin; prevent; small molecule; trafficking

DESCRIPTION (provided by applicant): Approximately 23.6 million Americans (7.8% of the population) have diabetes - most needing to produce more insulin because of insulin resistance, yet nearly all with pancreatic ß-cell dysfunction. In ß-cells, insulin synthesis begins with the precursor, preproinsulin, which must undergo co-translational translocation into the endoplasmic reticulum (ER), signal peptide (SP) cleavage, and downstream proinsulin folding. These earliest events are critical to insulin biosynthesis, but they are relatively understudied. Over the past three years, four preproinsulin SP mutations have been reported to cause human diabetes that make investigation of these earliest events especially timely. While it is known that insulin haploinsufficiency does not cause diabetes, patients with preproinsulin SP mutations are heterozygotes, suggesting that mutants act in a dominant- negative fashion. The molecular mechanisms of ß-cell failure caused by these mutants remain unknown. Interestingly, diabetes phenotypes associated with the SP mutants ranges from severe neonatal-onset diabetes caused by A(SP24)D, to mild adult-onset diabetes associated with R(SP6)C or H. I hypothesize that these two classes of SP mutants cause ß-cell failure through two distinct mechanisms. In one case, I propose that inefficient co-translational translocation [of R(SP6)C] causes cytosolic accumulation of untranslocated mutant that is slowly toxic to ß-cells, leading to adult-onset diabetes that may be akin to the pathogenesis of Alzheimer's and some other neurodegenerative diseases. In the other case, I propose that failed SP cleavage [of A(SP24)D] disturbs downstream proinsulin folding, causing ER retention of the mutant that abnormally interacts with co-expressed wild-type (WT) proinsulin and blocks its ER exit in trans, decreasing insulin production and initiating severe insulin-deficient diabetes in early life. This proposal aims to better understand the coordination of the earliest events of insulin biosynthesis and define the molecular mechanisms of ß-cell failure caused by defects of those events. The ultimate goal is to develop novel strategies to prevent development of diabetes caused by misfolded (pre)proinsulin. Three Specific Aims are proposed: 1) To examine coordination of preproinsulin co-translational translocation, SP cleavage, and downstream proinsulin folding; 2) To define genetic manipulations that would allow WT proinsulin to escape from blockade caused by mutant (pre)proinsulins; 3) To identify small molecules that could prevent ß-cell failure caused by mutant preproinsulins. Accumulating evidence suggests that ER stress and proinsulin misfolding plays a role in the pathogenesis of the most common form of diabetes (type 2) which does not involve any preproinsulin coding sequence mutations. These new diabetogenic mutants, in which underlying (pre)proinsulin mishandling-misfolding is unequivocal, are ideal models for understanding molecular mechanisms of ß-cell failure, and for testing experimental therapies aiming at preventing diabetes.

描述（由申请人提供）：大约2360万美国人（占人口的7.8%）患有糖尿病——大多数人由于胰岛素抵抗而需要分泌更多的胰岛素，但几乎所有人都患有胰腺ß-细胞功能障碍。在ß-细胞中，胰岛素合成始于前体胰岛素前原，胰岛素前原必须经过共翻译易位进入内质网（ER）、信号肽（SP）裂解和下游胰岛素前原折叠。这些最早的事件对胰岛素的生物合成至关重要，但它们的研究相对不足。在过去的三年中，已经报道了四种胰岛素前原SP突变导致人类糖尿病，这使得对这些早期事件的研究变得尤为及时。虽然已知胰岛素单倍体功能不全不会导致糖尿病，但具有胰岛素前原SP突变的患者是杂合子，这表明突变以显性负性方式起作用。由这些突变体引起的ß-细胞衰竭的分子机制尚不清楚。有趣的是，与SP突变体相关的糖尿病表型范围从由A(SP24)D引起的严重新生儿糖尿病，到与R(SP6)C或h相关的轻度成人糖尿病。我假设这两类SP突变体通过两种不同的机制引起ß-细胞衰竭。在一个案例中，我提出，[R(SP6)C]的低效共翻译易位导致未易位突变体的胞质积累，这些突变体对ß-细胞缓慢毒性，导致成人发病的糖尿病，这可能类似于阿尔茨海默病和其他一些神经退行性疾病的发病机制。在另一种情况下，我认为[A(SP24)D]的SP切割失败扰乱了下游的胰岛素原折叠，导致突变体的内质网保留，该突变体与共表达的野生型（WT）胰岛素原异常相互作用，并阻断其在反式中的内质网出口，减少胰岛素的产生，并在生命早期引发严重的胰岛素缺乏症糖尿病。该建议旨在更好地理解胰岛素生物合成最早事件的协调，并定义由这些事件缺陷引起的ß-细胞衰竭的分子机制。最终目标是开发新的策略来预防由错误折叠（前）胰岛素原引起的糖尿病的发展。提出了三个具体目的：1)研究胰岛素前原共翻译易位、SP切割和下游胰岛素原折叠的协调性；2)定义基因操作，使WT胰岛素原逃脱突变（预）胰岛素原引起的封锁；3)鉴定可以预防突变前胰岛素引起的ß-细胞衰竭的小分子。越来越多的证据表明，内质网应激和胰岛素原错误折叠在最常见的糖尿病（2型）的发病机制中起作用，而2型糖尿病不涉及任何胰岛素前原编码序列突变。这些新的致糖尿病突变，其中潜在的（前）胰岛素原错误处理-错误折叠是明确的，是理解ß-细胞衰竭的分子机制和测试旨在预防糖尿病的实验性疗法的理想模型。