Signaling mechanisms that detect stress and maintain homeostasis

检测压力和维持体内平衡的信号机制

• 批准号: 10219290
• 负责人: T Keith Blackwell
• 金额: $ 49.62万
• 依托单位: JOSLIN DIABETES CENTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-20 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10219290
• 关键词: 70-kDa Ribosomal Protein S6 Kinases; Acute; Affect; Aging; Area; CRISPR/Cas technology; Caenorhabditis elegans; Cell physiology; Data; Disease; Drug Metabolic Detoxication; Enzyme Activation; Event; Growth; Health; Homeostasis; Human; Human Development; Longevity; Mass Spectrum Analysis; Mediating; Metabolic; Mitochondria; Modeling; Modification; Molecular Chaperones; Nox enzyme; Organism; Orthologous Gene; Oxidation-Reduction; Phenotype; Phosphotransferases; Play; Proteins; Proto-Oncogene Proteins c-akt; ROCK1 gene; Reactive Oxygen Species; Regulation; Research; Role; Screening Result; Signal Transduction; Stress; Work; base; biological adaptation to stress; genome editing; in vivo; interest; p38 Mitogen Activated Protein Kinase; response; sensor; small molecule; stress reactivity; transcription factor

Project Summary This MIRA proposal focuses on two overlapping areas: stress response regulation and the functions of redox-based signaling in vivo. It is a fundamentally important problem how organisms detect and respond to different forms of stress. Much has been learned in this area but we still have a very incomplete understanding of how some stresses are detected, including reactive small molecules such as ROS. For many years my group has studied stress responses and aging in C. elegans, focusing on the Nrf2 transcription factor ortholog SKN-1. Nrf2 mediates a conserved detoxification response to reactive small molecules but has many additional functions, and is of great importance in health and disease. Working in C. elegans we have defined a number of aspects of SKN-1/Nrf2 regulation and functions, including its major role in longevity assurance. We have recently uncovered an exciting mechanism of SKN-1/Nrf regulation that forms the basis for this new research direction. We find that SKN-1 and human Nrf2 are activated at the ER by a localized ROS signal that can derive from the ER, NOX enzyme activation induced by stress, or mitochondria. This signal induces sulfenylation of a single Cys within the kinase activation loop of the ER unfolded protein sensor IRE-1, resulting in acute inhibition of the IRE-1 unfolded protein response and activation of p38 signaling at IRE-1 through a second sulfenylation event. p38 signaling in turn activates SKN-1/Nrf2. Remarkably, other kinases of major interest (AKT, p70S6K, ROCK1) seem to be regulated through sulfenylation of the same Cys. The data reveal an unexpected IRE-1 function that is regulated by a redox switch, a major stress sensor for SKN- 1/Nrf2, and a possible rationale for how redox stress can affect so many cellular processes. They also suggest that the scope and functional versatility of Cys-based signaling are much wider than is generally appreciated. In our proposed research we will continue to identify mechanisms of SKN-1/Nrf2 regulation and their functions in vivo, but will also cast our net wider in utilizing the advantages of C. elegans to explore mechanisms and functions of Cys redox signaling in the context of stress, growth, and other conditions. We will refine models for IRE-1 regulation of SKN-1/Nrf2 and its functions in vivo. We will also similarly study SKN- 1/Nrf2 regulation by the chaperone TRIC, another mechanism we have identified that may involve redox signaling, and build upon screening results to develop new models for SKN-1/Nrf2 regulation. Using mass spectrometry (MS), we will collaboratively identify C. elegans proteins that are Cys-sulfenylated under stress and growth conditions. We will investigate regulatory and in vivo implications of this modification for the kinases indicated above, and candidates chosen from our MS data. C. elegans will be ideal for this work because of the relative rapidity of Cas9/CRISPR genome editing, and phenotypic analyses. Our research will reveal stress-responsive regulatory mechanisms of fundamental interest, and take major steps towards identifying new mechanistic targets and functional implications of redox signaling in vivo.

项目摘要 MIRA的建议集中在两个重叠的领域：应激反应调节和 体内基于氧化还原的信号传导。生物体如何检测和响应 不同形式的压力。在这方面已经学到了很多东西，但我们仍然有一个非常不完整的理解 如何检测一些压力，包括活性小分子，如ROS。多年来，我 研究小组研究了C. elegans，专注于Nrf 2转录因子直系同源物 SKN-1。nrf 2介导对反应性小分子的保守解毒反应，但具有许多 它具有额外的功能，对健康和疾病非常重要。工作在C。我们已经定义了 SKN-1/Nrf 2调节和功能的许多方面，包括其在寿命保证中的主要作用。 我们最近发现了一种令人兴奋的SKN-1/Nrf调节机制，该机制构成了 这个新的研究方向。我们发现SKN-1和人Nrf 2在ER被局部ROS激活， 信号可以来自ER、应激诱导的NOX酶激活或线粒体。该信号 诱导ER未折叠蛋白传感器IRE-1的激酶活化环内的单个Cys的亚磺酰化， 导致IRE-1未折叠蛋白应答的急性抑制和IRE-1处p38信号传导的激活 通过第二次亚磺酰化事件。p38信号反过来激活SKN-1/Nrf 2。值得注意的是， 主要感兴趣的（AKT，p70 S6 K，ROCK 1）似乎通过相同Cys的亚磺酰化来调节。的 数据揭示了一种意想不到的IRE-1功能，它由氧化还原开关调节，氧化还原开关是SKN的主要应力传感器， 1/Nrf 2，以及氧化还原应激如何影响如此多的细胞过程的可能原理。他们还建议 基于Cys的信号传导的范围和功能多样性比通常理解的要宽得多。 在我们提出的研究中，我们将继续确定SKN-1/Nrf 2调节的机制及其作用机制。 功能，但也将我们的网络更广泛地利用C的优势。elegans探索 Cys氧化还原信号传导在应激、生长和其他条件下的机制和功能。我们 将完善IRE-1调节SKN-1/Nrf 2及其体内功能的模型。我们也将同样研究SKN- 1/Nrf 2受到伴侣TRIC的调节，这是我们发现的另一种可能涉及氧化还原的机制 信号转导，并建立在筛选结果，以开发新的模型SKN-1/Nrf 2的调控。利用大众 质谱（MS），我们将合作确定C。在应激下Cys-亚磺酰化的线虫蛋白质 和生长条件。我们将研究这种修饰对调节和体内的影响， 激酶，以及从我们的MS数据中选择的候选物。C. elegans将是这项工作的理想选择 因为Cas9/CRISPR基因组编辑和表型分析的相对快速。我们的研究将 揭示具有根本利益的压力响应调节机制，并采取重大步骤， 确定体内氧化还原信号传导的新机制靶点和功能意义。