Sreeganga S. Chandra, Ph.D. - US grants

DESCRIPTION (provided by applicant): Parkinson's disease (PD) is a progressive neurodegenerative disease affecting millions of people. Synaptic dysfunction is an early event in the pathogenesis of the disease occurring prior to the onset of symptoms. Therapeutic interventions at these early stages hold the promise of slowing or even halting the disease. The biggest obstacle to the development of such therapies is a lack of knowledge of early molecular and synaptic events that occur in PD. Therefore, our long term objective is to understand the mechanisms of synaptic dysfunction in PD. a-Synuclein was the first gene identified to cause dominant familial PD, and is also the major constituent of Lewy bodies, the pathological hallmarks of PD. Hence, considerable effort is being directed at understanding the role of a-synuclein in the pathogenesis of PD. However, these efforts are focused mostly on toxic gain-of- function approaches, such as its aggregation. In contrast, little is known about the normal physiological functions of a-synuclein at the presynaptic terminal and its contribution to PD. This is the objective of our proposal. We have generated a novel mouse that lacks all synucleins to facilitate these studies. To attain the objective of this application, three aims are pursued. First, the interaction of synucleins with newly identified physiological partners and the bilayer will be examined. Second, the synaptic transmission deficits of synuclein null neurons will be characterized in culture and in vivo. Third, we will study how PD mutations impact these physiological properties and functions. Achieving these goals is important for human health, given aging demographics and the increasing prevalence of neurodegenerative diseases. PUBLIC HEALTH RELEVANCE: a -Synuclein is a protein that forms clumps in the brains of Parkinson's patients. We are investigating what the normal function of a-synuclein is in the brain and whether alterations of this function plays a part in Parkinson's disease. This study will allow us, in time, to design early therapies for preventing cell death in Parkinson's disease.

? DESCRIPTION (provided by applicant): Palmitoylation is the covalent attachment of C16 fatty acids to proteins. Palmitoylation is a dynamic post- translational modification and unlike other stable lipid modifications (isoprenylation, farnesylation). One of the major enzymes that remove this lipid modification is palmitoyl protein thioesterase 1 (PPT1). Recessive loss-of-function mutations in PPT1 cause Neuronal Ceroid Lipofuscinosis (NCL), a progressive neurodegenerative disease with lysosomal pathology. Complete loss of PPT1 enzymatic activity leads to an infantile form of the disease, while residual activity (5-10%) leads to adult-onset NCL. Patients accumulate lipidated peptides suggesting that deficient depalmitoylation leads to compromised protein degradation. These clinical findings strongly suggest that PPT1 activity is critical for neuronal function and health. Despite its importance, the substrates of PPT1 remain to be defined. In this proposal, we aim to identify PPT1 substrates based on an unbiased proteomic screen comparing the palmitomes of wildtype and PPT1 knockout brains. Then, we will examine which of these substrates are depalmitoylated in response to neuronal activity. These experiments will delineate an important, yet understudied, aspect of neuronal cell biology and elucidate the mechanisms of NCL. Achieving these goals is important for human health, given the direct links between PPT1 and NCL as well as the wide range of neurodegenerative disorders that involve protein palmitoylation.

PROJECT SUMMARY Mutations in the GBA gene that encodes glucocerebrosidase 1 (GCase1) have been identified as the most common genetic risk factor for Parkinson's disease (PD) and Dementia with Lewy Bodies (DLB). Homozygous loss-of-function GBA mutations causes Gaucher's disease (GD). Notably, PD and DLB patients with GBA mutations exhibit ?-synuclein pathology strongly suggesting a link between GCase1 and ?-synuclein (gene:SNCA). However, the underlying mechanisms linking these two proteins and diseases remain unclear. GCase1 is a lysosomal enzyme which hydrolyzes glucosylceramide (GlcCer) to ceramide and glucose. GD patients accumulate not only the primary substrate GlcCer, but also downstream bioactive lipids such as glucosylsphingosine (GlcSph) due to alterations in glycosphingolipid homeostasis. We recently showed that GlcSph triggers the formation of highly self-templating ?-synuclein oligomers (Taguchi et al., 2017). Using new long-lived mouse models of GD crossed to ?-synuclein transgenics (Gba/SNCA mice), we showed that GlcSph levels increase in the brains of young mice, months prior to increases in GlcCer levels and that glycosphingolipid accumulations are co-localized with ?-synuclein pathology (Taguchi et al., 2017). Our hypothesis for the proposed research is that GlcSph is an initial driver of ?-synuclein pathology in GBA-linked PD and DLB. In this application, we will examine in detail the spatial and temporal profiles of glycosphingolipids and phospholipids in brains of Gba/SNCA mice. We will also test whether genetically altering GlcSph generating enzymes will ameliorate ?-synuclein pathology in Gba/SNCA mice as well as GBA-linked PD and DLB patient iPSC derived neurons. These studies will determine whether therapeutic reductions of elevated GlcSph is beneficial in GBA- linked PD and DLB. Achieving these goals is important for human health, given the massive personal and societal burden imposed by the elderly population suffering from PD and DLB.