Project Grant F31HL188940

HDAC7-DRIVEN REPROGRAMMING OF PKM2 FUNCTION IN LUNG VASCULAR PERMEABILITY - PROJECT SUMMARY ACUTE LUNG INJURY (ALI) IS CHARACTERIZED BY BREAKDOWN OF THE PULMONARY ENDOTHELIAL BARRIER, RESULTING IN VASCULAR LEAK, EDEMA, AND RESPIRATORY FAILURE. NO CURRENT THERAPIES DIRECTLY RESTORE ENDOTHELIAL INTEGRITY. PRELIMINARY DATA IDENTIFY THE CLASS IIA HISTONE DEACETYLASE HDAC7 AS A KEY DRIVER OF ENDOTHELIAL DYSFUNCTION. HDAC7 IS SELECTIVELY UPREGULATED IN HUMAN LUNG MICROVASCULAR ENDOTHELIAL CELLS EXPOSED TO LIPOPOLYSACCHARIDE (LPS), AND ENDOTHELIAL- SPECIFIC DELETION OF HDAC7 IN MICE MARKEDLY IMPROVES LUNG FUNCTION AND REDUCES VASCULAR INJURY. LPS INDUCES HDAC7 PHOSPHORYLATION AND NUCLEAR EXPORT, SUGGESTING A SHIFT TOWARD EXTRANUCLEAR SIGNALING ROLES DURING INFLAMMATORY STRESS. HDAC7 INTERACTS WITH PYRUVATE KINASE M2 (PKM2), A METABOLIC ENZYME THAT ADOPTS PRO- INFLAMMATORY SIGNALING FUNCTIONS WHEN IT TRANSITIONS FROM ITS GLYCOLYTIC TETRAMER TO ITS DIMERIC KINASE FORM. LPS ENHANCES HDAC7-PKM2 ASSOCIATION AND INCREASES PKM2 EXPRESSION AND PHOSPHORYLATION, WHILE PKM2 DEPLETION OR PHARMACOLOGIC STABILIZATION OF ITS TETRAMERIC STATE SIGNIFICANTLY ATTENUATES LPS-INDUCED ENDOTHELIAL BARRIER DISRUPTION. THESE DATA SUPPORT THE CENTRAL HYPOTHESIS THAT HDAC7 NUCLEAR EXPORT PROMOTES PKM2 REPROGRAMMING INTO A NUCLEAR, PRO-INFLAMMATORY EFFECTOR THAT DRIVES ENDOTHELIAL BARRIER FAILURE IN ALI. ALL KEY EXPERIMENTAL SYSTEMS, INCLUDING VALIDATED ENDOTHELIAL HDAC7 KNOCKOUT MICE, ESTABLISHED PERMEABILITY ASSAYS, AND PKM2-MODULATING TOOLS, ARE ALREADY IN PLACE, ENSURING FEASIBILITY FOR THE PROPOSED FELLOWSHIP WORK. THE MURINE STUDIES ARE STREAMLINED AROUND A SINGLE PRIMARY ENDPOINT WITH POWERED GROUP SIZES, AND MECHANISTIC ASSAYS RELY ON STANDARDIZED, ROUTINELY USED APPROACHES THAT MINIMIZE TECHNICAL RISK AND IMPROVE CLARITY OF INTERPRETATION. AIM 1 WILL DEFINE HOW HDAC7 NUCLEAR EXPORT REGULATES PKM2 LOCALIZATION, METABOLIC ACTIVITY, AND BARRIER FUNCTION IN VITRO AND IN VIVO. AIM 2 WILL DETERMINE THE NUCLEAR SIGNALING CONSEQUENCES OF HDAC7- DEPENDENT PKM2 IMPORT, FOCUSING ON STAT3 AND OTHER KEY INFLAMMATORY OUTPUTS USING TARGETED BIOCHEMICAL ANALYSES. BECAUSE ENDOTHELIAL BARRIER BREAKDOWN IS THE CENTRAL DRIVER OF RESPIRATORY FAILURE IN ALI AND ARDS, ELUCIDATING HOW HDAC7 REPROGRAMS PKM2 DURING INFLAMMATORY STRESS MAY IDENTIFY DRUGGABLE NODES CAPABLE OF RESTORING VASCULAR STABILITY. THE PROPOSED FELLOWSHIP PROVIDES INTEGRATED TRAINING IN MECHANISTIC BIOCHEMISTRY, ENDOTHELIAL BARRIER ANALYSES, AND IN VIVO LUNG INJURY MODELING, COUPLED WITH STRUCTURED GUIDANCE IN EXPERIMENTAL DESIGN, DATA INTERPRETATION, AND SCIENTIFIC COMMUNICATION. THIS ENVIRONMENT WILL EQUIP THE APPLICANT WITH THE CONCEPTUAL AND TECHNICAL FOUNDATION REQUIRED FOR AN INDEPENDENT RESEARCH CAREER FOCUSED ON VASCULAR BIOLOGY AND INFLAMMATORY LUNG DISEASE.