The interplay between the UPR and protein biogenesis at the ER

UPR 和 ER 蛋白质生物发生之间的相互作用

• 批准号: 10403561
• 负责人: MALAIYALAM MARIAPPAN
• 金额: $ 34.51万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10403561
• 关键词: ATP phosphohydrolase; Address; Antibodies; Apoptotic; Architecture; Attenuated; Beta Cell; Biochemical; Biogenesis; CRISPR/Cas technology; Carrier Proteins; Cell Death; Cells; Cessation of life; Chronic; Complex; Data; Defect; Degradation Pathway; Development; Diabetes Mellitus; Disease; Dominant-Negative Mutation; Electron Transport Complex III; Endoplasmic Reticulum; Enzymes; Funding; Genes; Homeostasis; Hormones; Human; Knowledge; Lead; Life; Link; Malignant Neoplasms; Mediating; Membrane Proteins; Messenger RNA; Molecular; Molecular Chaperones; Monitor; Mutation; Non-Insulin-Dependent Diabetes Mellitus; Pathway interactions; Phosphorylation; Phosphotransferases; Physiological; Play; Protein translocation; Proteins; Quality Control; RNA Splicing; Ribonucleases; Ribosomes; Role; Signal Transduction; Structure; Testing; Ubiquitination; Work; XBP1 gene; endoplasmic reticulum stress; human disease; insight; misfolded protein; novel; polycystic liver disease; polypeptide; prevent; protein folding; reconstitution; recruit; response; secretory protein; sensor; transcription factor

Project Summary/Abstract: Secretory and membrane proteins, which account for ~30% of all human proteins, are co-translationally translocated across or inserted into the endoplasmic reticulum (ER). These nascent polypeptides are folded into functional proteins with the help of chaperones and folding enzymes in the ER. Defects in protein folding lead to the accumulation of misfolded proteins and the triggering of ER stress, which activates the unfolded protein response (UPR). Of the three major UPR sensors, IRE1α is the most conserved ER-localized transmembrane kinase/RNase that is activated through oligomerization/phosphorylation upon ER stress. Once activated, IRE1α mediates the splicing of XBP1u mRNA to produce an active transcription factor, XBP1s, which drives expression of UPR target genes to mitigate ER stress. Also, IRE1α promiscuously cleaves ER- localized mRNAs through the regulated Ire1-dependent decay (RIDD) pathway to reduce the burden of the incoming protein load. Under chronic ER stress conditions, however, IRE1α switches from the pro-survival mode to pro-apoptotic mode, resulting in cell death, which is associated with human diseases including, type 2 diabetes and cancer. Despite the physiological importance, the factors that control activation and inactivation of IRE1α/XBP1 signaling remain unclear. We have recently discovered that IRE1α forms a complex with the Sec61/Sec63 translocon complex to access its mRNA substrates. In the current funding period, we have shown that the Sec61 translocon bridges IRE1α with the Sec63/BiP complex to turnoff IRE1α signaling during persistent ER stress. Our studies discovered that the Sec63/BiP complex is also responsible for freeing clogged Sec61 translocons as well as promoting protein folding in the ER. These new findings raise the hypothesis that the IRE1α/Sec61/Sec63 complex plays a central role in the activation and inactivation of IRE1α/XBP1 signaling to maintain ER homeostasis in cells. In the next funding period, we will test this hypothesis by (i) determining the role of this complex in making life-or-death decisions during ER stress; (ii) determining the architecture of the IRE1α/Sec61/Sec63/BiP complex; (iii) determining the role of this complex in sensing/responding to protein translocation defects in the ER. In an independent aim, we will establish a novel functional link between a cytosolic quality control and IRE1α/XBP1 signaling. We plan to use a combined approach of CRISPR/Cas9 edited cells, biochemical reconstitution, and structural approaches to address these problems. Overall, we expect these studies will provide a mechanistic insight into how the UPR and protein translocation/quality control pathways work together to maintain ER homeostasis. The knowledge gained from these studies will inform the development of possible treatments for several human diseases including diabetes, cancer, and polycystic liver diseases.

项目摘要/摘要： 秘书长和膜蛋白占所有人类蛋白的约30％，是共译的 易位或插入内质网（ER）。这些新生的多肽折叠 借助伴侣和ER中的折叠酶进入功能蛋白。蛋白质折叠的缺陷 导致错误折叠蛋白的积累和触发ER应力的触发，这激活了展开的 蛋白质反应（UPR）。在三个主要UPR传感器中，IRE1α是配置最多的ER定位 跨膜激酶/RNase通过在ER应力时通过低聚/磷酸化激活。一次 激活的IRE1α介导XBP1U mRNA的剪接以产生活性转录因子xbp1s， 这驱动UPR靶基因的表达以减轻急诊应力。此外，IRE1α混杂会切割Er- 通过受调节的IRE1依赖性衰减（RIDD）途径局部mRNA，以减少燃烧的燃烧 传入的蛋白质负荷。然而，在慢性ER应力条件下，IRE1α从促生寿使的转换 促凋亡模式的模式，导致细胞死亡，这与人类疾病有关，包括2型 糖尿病和癌症。尽管身体重要性，但控制激活和失活的因素 IRE1α/XBP1信号的传导仍不清楚。 我们最近发现，IRE1α与Sec61/sec63转运络合物形成一个复合物 访问其mRNA底物。在当前的资金期间，我们已经证明了SEC61转运桥 IRE1α具有SEC63/BIP复合物，可在持续性的ER应力期间进行IRE1α信号传导。我们的研究 发现SEC63/BIP综合体还负责释放Clegged Sec61 Cranslocons以及 促进ER中的蛋白质折叠。这些新发现提出了以下假设：IRE1α/sec61/sec63 复杂在IRE1α/XBP1信号的激活和失活以维持ER中起着核心作用 细胞中的稳态。在下一个资金期间，我们将通过（i）确定这一作用来检验这一假设 复杂的在ER压力期间做出生命或死亡决策； （ii）确定 IRE1α/sec61/sec63/bip复合物； （iii）确定该复合物在感测/反应蛋白质中的作用 ER中的易位缺陷。在一个独立的目标中，我们将建立一个新颖的功能联系 胞质质量控制和IRE1α/XBP1信号传导。我们计划使用CRISPR/CAS9的组合方法 编辑的细胞，生化重构以及解决这些问题的结构方法。总体而言，我们 期望这些研究将提供有关UPR和蛋白质易位/质量如何的机械洞察力 控制途径共同努力维护ER稳态。从这些研究中获得的知识将 告知开发几种人类疾病的可能治疗方法，包括糖尿病，癌症和 多囊肝病。