DCAF7/HDAC4/TFEB axis in acute lung injury

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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.