Project Grant R21NS135376

DNAJC13 P.N860S AS A PHYSIOLOGIC MODEL OFALPHA-SYNUCLEIN OVEREXPRESSION - PROJECT SUMMARY/ABSTRACT PARKINSON'S DISEASE (PD) IS A DEBILITATING NEURODEGENERATIVE DISORDER ASSOCIATED WITH THE LOSS OF DOPAMINE PRODUCING NEURONS AND THE ACCUMULATION OF PROTEIN AGGREGATES RICH IN A-SYNUCLEIN (LEWY BODIES). WHILE THE CAUSES OF PD ARE NOT FULLY KNOWN, RARE FAMILIES WITH MONOGENIC FORMS PROVIDE TRANSLATABLE TARGETS AND THE MODELS IN WHICH TO TEST NOVEL THERAPEUTIC STRATEGIES. FOR EXAMPLE, A-SYNUCLEIN (SNCA) WAS DISCOVERED TO BE THE MAJOR PROTEIN CONSTITUENT OF LEWY BODIES ONLY AFTER THE GENE ENCODING IT WAS LINKED TO FAMILIAL PD. OUR DISCOVERY OF SNCA MULTIPLICATIONS SUBSEQUENTLY PROVIDED THE RATIONALE AND MODELS TO DEVELOP MULTIPLE APPROACHES TO LOWER ASYN EXPRESSION THAT ARE CURRENTLY IN CLINICAL TRIALS. IN 2014, WE GENETICALLY LINKED A MISSENSE MUTATION (P.N855S) IN DNAJC13 (ENCODING RECEPTOR-MEDIATED ENDOCYTOSIS-8; RME-8) TO LATE-ONSET AUTOSOMAL-DOMINANT PARKINSONISM IN MULTI-INCIDENT MENNONITE KINDREDS (RME-8 PD). AFFECTED FAMILY MEMBERS HAD POST-MORTEM LEWY PATHOLOGY RICH IN A-SYNUCLEIN (ASYN) AND NEURONAL LOSS IN THE SUBSTANTIA NIGRA (SNPC). RME-8 IS A DNAJ-DOMAIN- CONTAINING PROTEIN THAT, TOGETHER WITH HEAT SHOCK COGNATE 70 (HSC70), STIMULATES: 1) THE REFOLDING OF MISFOLDED PROTEINS AND; 2) CLATHRIN REMOVAL FROM VESICLES. HENCE, LOSS OF RME-8 LEADS TO AN ACCUMULATION OF CLATHRIN COATED VESICLES, HYPER EXTENDED MEMBRANE TUBULES AND THE AGGREGATION OF CARGOS. GIVEN THIS BACKGROUND, WE DEVELOPED A DNAJC13 P. N860S KNOCK-IN (DKIHET; CORRESPONDING TO HUMAN P. N855S MUTATION) MOUSE MODEL. PRELIMINARY STUDIES IN DKIHET BRAIN AT 3 MONTHS OF AGE REVEAL THE RME-8 MUTATION: 1) INCREASES SOLUBLE MONOMERIC ASYN AND CLATHRIN COATED INTERMEDIATES; 2) CAUSES SYNAPTIC DELAY FOLLOWING REPETITIVE STIMULATION, AND; 3) IMPAIRS MOVEMENT INITIATION (RESULTING IN FREEZING) IN OPEN FIELD BEHAVIOR. THE PHENOTYPES DKIHET CONFERS ARE CONSISTENT WITH A PATHOLOGIC GAIN OF FUNCTION AND REMINISCENT OF A 'SYNAPTOPATHY. IN THIS PROPOSAL WE AIM TO MORE FULLY CHARACTERIZE THE DKIHET MODEL WITH RESPECT TO MUTANT AND WILDTYPE RME-8 PROTEIN-BINDING IN SYNAPTOSOMES FROM MICRODISSECTED STRIATUM. WE WILL MORE FULLY CHARACTERIZE DIFFERENCES IN ASYN BIOLOGY, INCLUDING PHOSPHORYLATED AND HIGHER MOLECULAR WEIGHT SPECIES. WE WILL ALSO LOOK AT THE MUTATION'S IMPACT ON DOPAMINERGIC FUNCTION, INCLUDING BIOCHEMICAL, PHYSIOLOGIC AND MORPHOLOGIC ASSESSMENTS. LASTLY, GIVEN RME-8 IS ALSO AN INTERFERON- RESPONSIVE GENE WE WILL EXPLORE NEUROIMMUNE BIOLOGY AND FOCUS ON MICROGLIAL PRUNING OF DOPAMINERGIC TERMINALS. COMPLETION OF THESE EXPERIMENTS WILL DEFINE THE BIOLOGIC ROLE OF DNAJC13 P.N855S, HIGHLIGHT ITS MECHANISM OF ACTION IN DISEASE PATHOGENESIS, AND DEMONSTRATE THE UTILITY OF DKIHET MICE.