Project Grant R21EY035812

CHARACTERIZATION OF THE NASCENT RETINAL PROTEOME REGULATING HIPPO SIGNALING DURING DAMAGE - PROJECT SUMMARY/ABSTRACT RETINAL DEGENERATIVE DISEASES SUCH AS AGE-RELATED MACULAR DEGENERATION AND GLAUCOMA, AS WELL AS TRAUMATIC INJURY, TYPICALLY LEAD TO LOSS OF RETINAL NEURONS AND THUS SIGHT. THEREFORE, THERE IS A CRITICAL NEED TO DEVELOP THERAPIES TO REGENERATE LOST RETINAL NEURONS FOR VISION RESTORATION. THE ABILITY FOR THIS TYPE OF INTRINSIC SELF-REPAIR NATURALLY OCCURS IN THE ZEBRAFISH RETINA. HERE, IN RESPONSE TO RETINAL DAMAGE, THE MULLER GLIAL CELLS (MGS) REPROGRAM TO A PROLIFERATIVE, PROGENITOR STATE CAPABLE OF GIVING RISE TO NEW RETINAL NEURONS THAT INTEGRATE INTO THE PRE-EXISTING RETINAL CIRCUITRY AND RESTORE VISION. WHILE MAMMALIAN MGS DO NOT EXHIBIT THE REGENERATIVE POTENTIAL OF THE ZEBRAFISH, THEY HAVE SIMILAR GENE EXPRESSION AND HOMEOSTATIC ROLES. ADDITIONALLY, IN RESPONSE TO DAMAGE, MOUSE MGS DISPLAY VERY LIMITED AND TRANSIENT ENTRY INTO THE CELL CYCLE. THESE DATA SUGGEST THAT AN INTRINSIC PROLIFERATIVE AND/OR REGENERATIVE BLOCK LIKELY PREVENTS MAMMALIAN MGS FROM UNDERGOING SUSTAINED CELL CYCLE RE-ENTRY AND ACQUIRING A MULTIPOTENTIAL PROGENITOR- LIKE STATE. RECENTLY, IT WAS SHOWN THAT THE HIPPO SIGNALING PATHWAY FUNCTIONS TO REPRESS SUSTAINED ACTIVITY OF THE TEAD TRANSCRIPTION COFACTOR YAP, WHICH IS REQUIRED FOR MG PROLIFERATION AND REPROGRAMMING. BY MANIPULATING THE HIPPO PATHWAY, MAMMALIAN MGS MIGHT BE ABLE TO ELICIT A ROBUST REGENERATIVE RESPONSE ANALOGOUS TO THE ZEBRAFISH. HOWEVER, THE UPSTREAM COMPONENTS THAT TRIGGER INITIAL YAP ACTIVATION IN MGS AND SUBSEQUENT REPRESSION BY HIPPO ARE STILL COMPLETELY UNKNOWN THEREBY LIMITING MOLECULAR ENTRY POINTS FOR THERAPEUTIC INTERVENTION. TO COMPLICATE MATTERS, THE HIPPO PATHWAY RESPONDS TO A WIDE VARIETY OF CONTEXT-DEPENDENT BIOCHEMICAL AND BIOPHYSICAL INPUTS. TO MOVE THE FIELD FORWARD WITH AN EYE TOWARD CLINICAL TRANSLATION, WE MUST IDENTIFY THE MG-SPECIFIC REGULATORS OF YAP AND THE HIPPO PATHWAY. THEREFORE, THIS PROPOSAL AIMS TO USE CELL TYPE-SPECIFIC METABOLIC LABELING AND PURIFICATION OF PROTEINS FOLLOWED BY MASS SPECTROSCOPY ON DAMAGED MOUSE RETINAE. THIS APPROACH, WHICH HAS YET TO BE PERFORMED ON THE MAMMALIAN RETINA, WILL ALLOW US TO DECIPHER THE NASCENT PROTEOMES OF DAMAGED PHOTORECEPTORS AND GANGLION CELLS ALONG WITH THE IMMEDIATE MG RESPONSE DURING THE PERIOD OF YAP ACTIVATION AND SUBSEQUENT HIPPO SIGNALING. THIS INFORMATION WILL GENERATE NEW HYPOTHESES AND LAY THE FOUNDATION FOR FUTURE STUDIES AIMED AT BROADENING POTENTIAL THERAPEUTIC TARGETS FOR MODULATION OF THE HIPPO PATHWAY TO PROMOTE RETINAL REGENERATION.