Degradation of Endoplasmic Reticulum Chaperones by Chaperone-mediated Autophagy

伴侣介导的自噬对内质网伴侣的降解

• 批准号: 7675799
• 负责人: Roberta Kiffin
• 金额: $ 4.62万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7675799
• 关键词: Affect; Age; Aging; Amino Acids; Autophagocytosis; Biochemical; Biological Assay; Cell Survival; Cells; Characteristics; Cultured Cells; Data; Degradation Pathway; Diabetes Mellitus; Disease; Endoplasmic Reticulum; Ensure; Excision; Failure; GRP94; Goals; Homeostasis; Human; Image; Laboratories; Lead; Liver; Liver diseases; Living Wills; Lysosomes; Maintenance; Mediating; Metabolic Diseases; Modeling; Molecular; Molecular Chaperones; Nerve Degeneration; Neurodegenerative Disorders; Nutrient; Nutritional; Organelles; Organism; Pathogenesis; Pathway interactions; Play; Protein Biosynthesis; Proteins; Proteome; Quality Control; Recovery; Regulation; Rodent; Role; Source; Starvation; Stress; System; Testing; Tissues; Transgenic Mice; Ubiquitin; Up-Regulation; base; calreticulin; coping; deprivation; endoplasmic reticulum stress; genetic manipulation; multicatalytic endopeptidase complex; novel; novel strategies; prevent; protein folding; public health relevance; response; senescence

DESCRIPTION (provided by applicant): Autophagy is a major mechanism for cellular quality control, as it is responsible for the elimination of altered intracellular components in lysosomes. Autophagy also contributes to the maintenance of cellular protein and organelle homeostasis. Among these organelles, the endoplasmic reticulum (ER) plays a central role in protein biosynthesis and undergoes dramatic changes to accommodate the cellular demands of particular proteins. Chaperones resident in the ER are critical to assure proper folding of many newly synthesized proteins. Tight synthetic regulation of these chaperones ensures cell survival under a variety of both stress and non-stress conditions. Thus, increased synthesis of these chaperones is one of the first cellular responses to compromised protein folding in the ER, and the mechanisms leading to their upregulation are now well-characterized. However, comparatively little is known about the normal turnover of these chaperones or their eventual fate following their stress-induced translational upregulation. We have recently found that a selective type of autophagy, known as chaperone-mediated autophagy (CMA), may play a central role in the turnover of ER chaperones and in the recovery of normal ER homeostasis after stress. The purpose of this project is to elucidate the mechanism(s) of degradation of ER chaperones with special emphasis on the role of chaperone mediated autophagy (CMA) in their turnover. We will: 1) Examine the mechanisms for degradation of specific ER chaperones (GRP94, BiP and calreticulin) under normal basal conditions; 2) Determine if these mechanisms of degradation change in response to two stress conditions (ER stress and starvation), and 3) examine whether the previously described decline in CMA activity with age affects the degradation of ER chaperones and could contribute to the poor cellular response to ER stress in aging. For this purpose we will use both biochemical and image-based assays developed in our laboratory to track the degradation of ER chaperones in lysosomes by CMA. Furthermore, using genetic manipulations of critical components of CMA, both in cells in culture and in different tissues in rodents we will determine the consequences of failure in proper turnover of ER chaperones. Public Health Relevance: Inadequate cellular adaptation to ER stress has been identified as the pathogenetic basis of a growing list of devastating human disorders such as neurodegenerative disorders, metabolic diseases such as diabetes and severe liver diseases, among others. Consequently, a better understanding of the molecular mechanisms involved in the cellular response to ER stress could lead to novel approaches to modulate this response and to prevent its failure.

描述（由申请人提供）：自噬是细胞质量控制的主要机制，因为它负责消除溶酶体中改变的细胞内成分。自噬还有助于维持细胞蛋白质和细胞器稳态。在这些细胞器中，内质网 (ER) 在蛋白质生物合成中发挥着核心作用，并经历巨大的变化以适应特定蛋白质的细胞需求。内质网中的伴侣对于确保许多新合成蛋白质的正确折叠至关重要。这些分子伴侣的严格合成调节可确保细胞在各种应激和非应激条件下存活。因此，这些伴侣蛋白合成的增加是细胞对内质网蛋白质折叠受损的最初反应之一，导致其上调的机制现已得到充分表征。然而，人们对这些伴侣的正常更新或应激诱导的翻译上调后的最终命运知之甚少。我们最近发现，一种选择性类型的自噬，称为伴侣介导的自噬（CMA），可能在应激后内质网伴侣的更新和正常内质网稳态的恢复中发挥核心作用。该项目的目的是阐明 ER 伴侣的降解机制，特别强调伴侣介导的自噬 (CMA) 在其周转中的作用。我们将： 1) 检查正常基础条件下特定 ER 伴侣（GRP94、BiP 和钙网蛋白）的降解机制； 2) 确定这些降解机制是否会因两种应激条件（ER 应激和饥饿）而发生变化，3) 检查先前描述的 CMA 活性随年龄的下降是否会影响 ER 伴侣的降解，并可能导致衰老过程中细胞对 ER 应激的不良反应。为此，我们将使用我们实验室开发的生化和基于图像的测定法来跟踪 CMA 溶酶体中 ER 伴侣的降解。此外，通过对培养细胞和啮齿动物不同组织中的 CMA 关键成分进行遗传操作，我们将确定 ER 伴侣正常周转失败的后果。公共健康相关性：细胞对内质网应激的适应不足已被确定为越来越多的毁灭性人类疾病的发病基础，这些疾病包括神经退行性疾病、糖尿病等代谢疾病和严重的肝脏疾病等。因此，更好地了解细胞对内质网应激反应的分子机制可能会产生新的方法来调节这种反应并防止其失败。