Investigating the role of HSC70 AMPylation on nuclear localization and chaperone function

研究 HSC70 AMPylation 对核定位和伴侣功能的作用

• 批准号: 10490968
• 负责人: Shannon Marie Lacy
• 金额: $ 3.87万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10490968
• 关键词: ATP phosphohydrolase; Acute; Adenosine Monophosphate; Age; Amyloid fibers; Autophagocytosis; Autophagosome; Binding; Biological Assay; Caenorhabditis elegans; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Cause of Death; Cell Culture Techniques; Cell Fractionation; Cell Line; Cell Nucleus; Cell physiology; Cells; Cellular Stress; Cessation of life; Coronary heart disease; Cytosol; Data; Defect; Defense Mechanisms; Dysmyelopoietic Syndromes; Failure; Family; Goals; Heart; Heart Diseases; Heat Stress Disorders; Heat shock proteins; Heat-Shock Proteins 70; Heat-Shock Response; Hypoxia; Immunofluorescence Microscopy; Impairment; In Vitro; Influenza; Ischemia; Knock-out; Knockout Mice; Knowledge; Link; Luciferases; Mediating; Modeling; Molecular; Molecular Chaperones; Movement; Mutate; Myocardial Ischemia; Nuclear; Nuclear Import; Nuclear Localization Signal; Nuclear Protein; Nuclear Translocation; Orthologous Gene; Patients; Pharmacologic Substance; Physiology; Post-Translational Protein Processing; Process; Protein S; Proteins; Quality Control; Recycling; Regulation; Risk; Role; Signal Transduction; Site; Stress; System; Testing; Therapeutic; Threonine; Transgenic Organisms; Work; base; cardiovascular risk factor; chaperonin; heart function; misfolded protein; novel; nucleocytoplasmic transport; prevent; protein aggregation; protein degradation; protein folding; proteostasis; recruit; response

Project Summary Coronary heart disease, also called Ischemic Heart Disease (IHD), is the leading cause of death worldwide. Repeated episodes of cardiac stress in IHD patients leads to an increase in misfolded proteins in their hearts. In response to this acute onset of cellular stress, heat shock proteins (HSP)s are upregulated in myocardiocytes and help restore protein homeostasis (proteostasis). Heat shock cognate 71 kDa (HSC70) is an essential, HSP70 chaperone that is located in the cytosol in the absence of cellular stress. This ATP-dependent chaperone actively refolds, degrades, and disaggregates proteins. HSC70 is also critical for autophagy, a cellular recycling/degradation process that protects the heart from misfolded proteins following ischemia. Impaired HSC70 function has been implicated in heart disease, yet despite the critical role of HSC70 in cellular physiology, the molecular mechanisms that regulate nuclear transport and govern the disparate functions of HSC70 remain unknown. Protein AMPylation recently emerged as a distinct PTM that regulates HSP70 family chaperones through the addition of adenosine monophosphate (AMP) to threonine residues. This process is catalyzed by the mammalian AMPylase, HYPE. My preliminary work revealed that HYPE AMPylates HSC70 on two sites, and increased cellular AMPylation levels prevent the nuclear-cytosolic shuttling of HSC70 as well as its ATPase activity. I also found that, similar to decreased levels of the HSC70 ortholog HSP-1, increased AMPylation inhibits autophagosome formation in C. elegans. These data suggest that HYPE-mediated AMPylation of HSC70 inhibits its critical functions associated with maintaining proteostasis, including nuclear-cytosolic shuttling, protein folding, and autophagy. The goals of this proposal are to 1) define the functional impact of HYPE-mediated AMPylation on HSC70 and 2) determine the impact of AMPylation on HSC70’s nuclear-cytosolic shuttling mechanism. Ultimately, we aim to define how a novel post-translational modification (PTM), AMPylation, regulates HSC70 function in proteostasis and cardiovascular diseases. Advancing our knowledge of the mechanisms that control HSC70 function is critical to identify novel avenues to therapeutic strategies that capitalize on restoring HSC70 regulation.

项目摘要 冠心病，也称为缺血性心脏病（IHD），是全球死亡的主要原因。 IHD患者心脏应激的反复发作导致其心脏中错误折叠蛋白质的增加。在 心肌细胞对这种急性细胞应激的反应是热休克蛋白（HSP）的上调 并帮助恢复蛋白质稳态（proteostasis）。热休克同源物71 kDa（HSC 70）是一种必需的， 在没有细胞应激的情况下位于胞质溶胶中的HSP 70分子伴侣。这种依赖ATP的伴侣蛋白 主动地重折叠、降解和解聚蛋白质。HSC 70对自噬也是至关重要的，自噬是一种细胞自噬。 循环/降解过程，保护心脏免受缺血后错误折叠的蛋白质的影响。受损 HSC 70的功能与心脏病有关，尽管HSC 70在细胞生理学中起着关键作用， 调节核转运和支配HSC 70不同功能的分子机制仍然存在 未知 蛋白质AMP化最近作为一种独特的PTM出现，其通过调节HSP 70家族分子伴侣， 将腺苷一磷酸（AMP）添加至苏氨酸残基。这个过程是由哺乳动物 淀粉酶，HYPE。我的初步工作表明，HYPE在两个位点上使HSC 70发生AMPylates， 细胞AMP化水平阻止HSC 70的核-胞质穿梭以及其ATP酶活性。我也 发现，类似于HSC 70同源HSP-1水平的降低，AMP化的增加抑制了HSC 70同源HSP-1的表达。 自噬体形成。优雅的这些数据表明HYPE介导的HSC 70的AMP化抑制了HSC 70的表达。 其关键功能与维持蛋白质稳态有关，包括核-胞质穿梭，蛋白质 折叠和自噬。 本提案的目标是：1）确定HYPE介导的AMPylation对HSC 70的功能影响， 2)确定AMP化对HSC 70的核-胞质穿梭机制的影响。最终，我们的目标是 定义一种新的翻译后修饰（PTM），AMPylation，如何调节HSC 70在蛋白质稳定中的功能 和心血管疾病。推进我们对控制HSC 70功能的机制的了解至关重要 以确定利用恢复HSC 70调节的治疗策略的新途径。