DCAF7/HDAC4/TFEB axis in acute lung injury

DCAF7/HDAC4/TFEB 轴在急性肺损伤中的作用

• 批准号: 10183301
• 负责人: Yuan Liu
• 金额: $ 47.17万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10183301
• 关键词: Acute; Acute Lung Injury; Adrenal Cortex Hormones; Adult Respiratory Distress Syndrome; Alveolar; Attenuated; Autophagocytosis; Bacteria; Bacterial Infections; Behavior; Bioenergetics; Biogenesis; Cell Death; Cell Nucleus; Cells; Data; Defect; Disease; Dose; Energy Supply; Epithelial; Epithelial Cells; Equilibrium; Exhibits; FDA approved; Functional disorder; HDAC4 gene; Half-Life; Histone Deacetylase Inhibitor; Homeostasis; Housekeeping; Hypoxemia; Immune; Immunity; Impairment; Inflammasome; Inflammation; Inflammatory; Inflammatory Response; Injury; Innate Immune System; Libraries; Link; Lung; Lung Inflammation; Lysosomes; Mass Spectrum Analysis; Mechanical ventilation; Mediating; Mitochondria; Mitochondrial DNA; Modality; Modeling; Molecular; Natural Immunity; Oxygen; Oxygen Consumption; Pathway interactions; Patients; Pattern; Peptide Hydrolases; Permeability; Pharmaceutical Preparations; Pneumonia; Process; Proteins; Proteomics; Reactive Oxygen Species; Resolution; Role; Scheme; Severities; Signal Transduction; Stress; Structure of parenchyma of lung; Testing; Therapeutic; Transcription Coactivator; Work; airway epithelium; alveolar epithelium; base; cytokine; cytokine release syndrome; improved; inhibitor/antagonist; knock-down; mitochondrial dysfunction; molecular modeling; mortality; new therapeutic target; novel; overexpression; pathogen; preservation; prevent; protein degradation; receptor; recruit; screening; small molecule; targeted treatment; therapeutic development; transcription factor; ubiquitin-protein ligase

ABSTRACT Acute Respiratory Distress Syndrome (ARDS) or Acute Lung Injury (ALI) is an acute lung inflammatory process that is associated with an overall mortality ranging from 35% to 50%, and pneumonia is one of the most frequent causes. The central pathophysiology of ARDS/ALI is injury to the epithelium in the airway/alveoli, and overproduction of inflammatory factors in lung tissue. Mitochondria provide the energy supply for numerous cellular activities, and are also the powerhouses of immunity. Bacterial infection impairs mitochondria, and dysfunctional mitochondria need to be cleared through lysosomal degradation, a process termed autophagy/mitophagy. Endogenous damage-associated molecular patterns DAMPs (mtDNA, ATP, ROS) released from damaged mitochondria activate inflammasomes and inflammation. We identify that the deficiency of TFEB, a master transcription factor of autophagy and lysosome biogenesis, impairs mitophagy leading to deleterious DAMP and pro-inflammatory cytokines release, thus activating inflammasomes and inflammation. Through screening a library of FDA-approved drugs (1068 drugs), we identify that the HDAC inhibitor Panobinostat elevates TFEB protein abundance by inhibiting HDAC4. Through unbiased proteomic mass spectrometry analysis, we identify DCAF7 as a substrate receptor for CRL4 ubiquitin E3 ligase recruiting TFEB. DCAF7 overexpression dose-dependently decreases TFEB protein abundance. DCAF7 knockdown prolongs TFEB protein half-life and accumulates its protein levels in the nucleus. Specifically, our preliminary data suggest that 1) bacterial infection decreases TFEB protein levels; 2) TFEB deficiency aggravates harmful DAMP and cytokine release that in turn postpone inflammation resolution; 3) Panobinostat-mediated HDAC4 inhibition preserves TFEB proteins and alleviates bacteria-induced lung inflammation; 4) CRL4 ubiquitin E3 ligase subunit DCAF7 recruits TFEB for proteasomal degradation; 5) a novel small molecule DCAF7 inhibitor BC1753 protects TFEB against degradation. These data led to our hypothesis that inhibiting DCAF7/HDAC4 axis to prevent TFEB from degradation will attenuate bacterial infection-induced lung inflammation and improve inflammation resolution. We will conduct mechanistic studies to determine if TFEB exerts an essential role in bacterial lung inflammation through inhibiting necroptosis (Aim 1). We will examine the role of HDAC4 in regulating TFEB protein stability and lung inflammatory responses in experimental ALI models (Aim 2). We will also test if the small molecule DCAF7 inhibitor alleviates bacterial lung inflammation through preventing TFEB protein degradation (Aim 3). This will be the first study to intervene DCAF7/HDAC4/TFEB axis for inflammatory disease such as ARDS/ALI. Execution of this project will lay the groundwork for a fundamental, paradigm-changing therapeutic advance to regulate innate immunity and treat lung inflammation that will ultimately set the stage for a new translational initiative.

摘要 急性呼吸窘迫综合征(ARDS)或急性肺损伤(ALI)是一种急性肺 炎症过程，与总死亡率从35%到50%有关，以及 肺炎是最常见的原因之一。ARDS/ALI的中枢病理生理学机制是 呼吸道/肺泡上皮损伤和肺炎性因子过度产生 组织。线粒体为许多细胞活动提供能量供应，也是 豁免权的强国。细菌感染损害线粒体，并导致功能障碍 线粒体需要通过溶酶体降解来清除，这个过程被称为 自噬/有丝分裂。内源性损伤相关分子模式抑制(mtDNA， 损伤线粒体释放的三磷酸腺苷(ATP，ROS)激活炎症体和炎症。 我们发现，TFEB是自噬的主要转录因子，它的缺陷是 溶酶体生物发生，损害有丝分裂吞噬导致有害的湿和促炎 细胞因子释放，从而激活炎症体和炎症。通过筛选一名 FDA批准的药物库(1068种药物)，我们确定HDAC抑制剂帕诺比坦 通过抑制HDAC4提高TFEB蛋白丰度。通过无偏见的蛋白质组质量 光谱分析，我们确定DCAF7是CRL4泛素E3连接酶的底物受体 招募TFEB。DCAF7过表达可剂量依赖性地降低TFEB蛋白丰度。 DCAF7基因敲除延长了TFEB蛋白的半衰期，并使其蛋白水平在 原子核。具体地说，我们的初步数据表明，1)细菌感染降低了TFEB 蛋白质水平；2)TFEB缺乏加重了有害的湿气和细胞因子的释放，进而 推迟炎症消退；3)Panobinostat介导的HDAC4抑制保留TFEB 蛋白和减轻细菌引起的肺部炎症；4)CRL4泛素E3连接酶亚单位 DCAF7招募TFEB用于蛋白酶体降解；5)一种新的小分子DCAF7抑制剂 BC1753保护TFEB免受降解。这些数据导致了我们的假设，抑制 DCAF7/HDAC4轴防止TFEB降解将减弱细菌感染诱导 改善肺部炎症，改善炎症消退。我们将进行机理研究，以 确定TFEB是否通过抑制细菌性肺部炎症发挥重要作用 坏死性下垂(目标1)。我们将研究HDAC4在调节TFEB蛋白稳定性中的作用 和实验性ALI模型的肺炎性反应(目标2)。我们还将测试是否 小分子DCAF7抑制剂通过预防TFEB减轻细菌性肺部炎症 蛋白质降解(目标3)。这将是第一个干预DCAF7/HDAC4/TFEB轴的研究 用于炎症性疾病，如ARDS/ALI。该项目的实施将为 调节先天免疫和治疗的根本性、改变范式的治疗进展 肺部炎症，最终将为一项新的翻译倡议奠定基础。