Functional analyses of ER stress sensor proteins

内质网应激传感器蛋白的功能分析

• 批准号: 13480239
• 负责人: KOHNO Kenji
• 金额: $ 9.54万
• 依托单位: NARA INSTITUTE OF SCIENCE AND TECHNOLOGY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13480239/
• 关键词: ER stress; Unfolded Protein Response; IRE1; BiP; transport vesicle; ストレスセンサー; マイクロアレイ; 28srRNA; ER stress; Unfolded Protein Response; IRE1; BiP; transport vesicle; ストレスセンサー; マイクロアレイ; 28srRNA

1. The accumulation of unfolded proteins in the endoplasmic reticulum (ER) induces a transcriptional response in the nucleus, leading to the upregulation of many ER-resident proteins involved in protein folding, to protect cells from ER stress. This response is called the unfolded protein response (UPR), which is essential for maintaining the ER homeostasis. Two important factors for UPR have been identified. One is an ER stress sensor protein Ire1, and the other is a transcription factor Hac1. Overexpression of IRE1 or HAC1 specifically suppressed growth defects of sec mutants defective in COPII vesicle formation. These findings suggest that the activation of the UPR affects ER-to-Golgi transport via stimulation of COPII vesicle formation from the ER.Induction of UPR needs the activation of Ire1. It was previously hypothesized that BiP/Kar2 plays a direct role in the signaling mechanism. In this model, association of BiP/Kar2 with Ire1 represses the UPR pathway while under conditions of ER stress, BiP/Kar2 dissociation leads to activation. To test this model, we analyzed five temperature-sensitive alleles of the yeast KAR2 gene. When cells carrying a mutation in the Kar2 substrate-binding domain were incubated at the restrictive temperature, association of Kar2 to Ire1 was disrupted, and the UPR pathway was activated even in the absence of extrinsic ER stress. Conversely, cells carrying a mutation in the Kar2 ATPase domain, in which Kar2 poorly dissociated from Ire1 even in the presence of tunicamycin, a potent inducer of ER stress, were unable to activate the pathway. Our findings provide strong evidence in support of BiP/Kar2-dependent Ire1 regulation model and suggest that Ire1 associate with Kar2 as a chaperone substrate.

1。内质网（ER）中展开的蛋白的积累诱导细胞核中的转录反应，从而导致许多参与蛋白质折叠的ER居民蛋白的上调，以保护细胞免受ER应激。这种反应称为展开的蛋白质反应（UPR），这对于维持ER稳态至关重要。已经确定了两个重要因素。一个是ER应力传感器蛋白IRE1，另一个是转录因子HAC1。 IRE1或HAC1的过表达特异性抑制了Copii囊泡形成中有缺陷的SEC突变体的生长缺陷。这些发现表明，UPR的激活会通过刺激ER诱导UPR诱导的copi囊泡形成来影响ER到高尔基体的运输。以前假设BIP/KAR2在信号传导机理中起着直接作用。在此模型中，在ER应力条件下，BIP/KAR2与IRE1的关联抑制了UPR途径，BIP/KAR2解离会导致激活。为了测试该模型，我们分析了酵母KAR2基因的五个温度敏感性等位基因。当在限制温度下孵育Kar2底物结合结构域中的突变的细胞时，Kar2与IRE1的关联被破坏，即使在没有外部ER应力的情况下，UPR途径也被激活。相反，在KAR2 ATPase结构域中携带突变的细胞，即即使在有有效的ER应力诱导剂的有效诱导剂的情况下，KAR2与IRE1差的细胞也无法激活该途径。我们的发现提供了支持BIP/KAR2依赖性IRE1调节模型的有力证据，并表明IRE1将Kar2与伴侣蛋白底物相关。