Kelvin Luk - US grants

? DESCRIPTION (provided by applicant): The accumulation of misfolded proteins represents a common pathological mechanism of most major neurodegenerative disorders. Neuronal inclusions comprised of aggregated a-Synuclein (aSyn) are known as Lewy bodies (LBs) and Lewy-neurites (LNs), and represent a key histopathological feature of Parkinson's disease (PD), and a family of related disorders known as synucleinopathies that affect as many as 1 million individuals in the U.S. alone. Mutations in the SNCA gene encoding aSyn also cause familial PD but while histological and genetic evidence firmly indicate a correlation between aSyn accumulation and disease, how aSyn pathology forms and whether it directly contributes to disease remains unclear. Abnormal aSyn catalyzes the misfolding of the normal protein and it has recently been demonstrated that minute quantities of aSyn aggregates can trigger the formation of toxic LBs/LNs in cultured neurons. Misfolded aSyn also induces the formation of LBs/LNs in healthy non-transgenic mice. In both human PD and animal models, aSyn pathology progressively propagates and spreads to neuroanatomically connected regions, reminiscent of prion diseases. Importantly, animals with LBs/LNs recapitulate the cardinal features of PD, including progressive loss of dopamine-producing neurons and locomotor deficits. This proposed research plan addresses key biological questions posed by these findings, and combines novel in vitro, cell-based, and in vivo tools to further understand how LBs/LNs are form, propagate, and ultimately contribute to neurodegeneration and neurological symptoms. Aim 1 examines whether neurons in multiple brain regions develop LBs/LNs following inoculation with misfolded recombinant aSyn and subsequently undergo cell death. A recently developed tissue processing method will be used to determine if LBs/LNs spread via neuronal projections, as hypothesized for human PD, or by other mechanisms. Behavioral tests will then reveal if specific clinicopathological correlations exist. Aim 2 will define the molecular interactions that govern how abnormal aSyn triggers the conversion of normal aSyn in LBs/LNs, by testing the ability of mutant aSyn sequences to seed pathology in both cells and in vivo following stereotactic injection. Finally, Aim 3 will elucidate the cellular and molecular mechanisms by which aSyn induce intracellular pathology by using cell-based, in vivo, and proteomics approaches to compare LB/LN-inducing and non-inducing aSyn mutants that we have recently discovered. Completion of these studies should provide valuable insights into the potential mechanisms by which aSyn contribute to the progression of PD. Increased understanding of the pathogenesis of this and related synucleinopathies should ultimately result in disease-modifying therapies for this group of incurable disorders.

The accumulation of misfolded proteins represents a common pathological mechanism of most major neurodegenerative disorders. Neuronal inclusions comprised of aggregated ?-Synuclein (aSyn), known as Lewy bodies (LBs) and Lewy-neurites (LNs), represent a key histopathological feature of Parkinson's disease (PD) and a family of related disorders known as synucleinopathies, most notably Dementia with LBs (DLB). LBs are also a prominent feature in nearly half of Alzheimer's disease subjects. Mutations and amplifications in the SNCA gene encoding aSyn also cause familial forms of PD. Although a large body of histological and genetic evidence firmly indicate a correlation between aSyn accumulation and disease, it remains unclear how aSyn pathology actually forms and subsequently contributes to disease. We and others recently demonstrated that minute quantities of recombinant or patient-derived aSyn aggregates can catalyze the formation of toxic LBs/LNs in cultured neurons and healthy non-transgenic mice. In both human PD and animal models, this ?seeded? aSyn pathology progressively propagates and spreads to neuroanatomically connected regions, reminiscent of prion diseases. Importantly, animals with LBs/LNs recapitulate the cardinal features of PD, including progressive loss of dopamine-producing neurons and locomotor deficits. This R01 renewal addresses several key biological questions posed by our earlier findings and combines novel molecular, in vivo, and computational tools to further understand how LBs/LNs form, propagate, and ultimately contribute to neurodegeneration and neurological symptoms. Aim 1 will identify at the neuron subtypes that develop LBs/LNs following inoculation with misfolded aSyn. By combining traditional histological methods with FACS-assisted single-neuron RNAseq, we will determine the molecular signatures associated with subpopulations that are vulnerable or resistant to LBs/LNs formation. Aim 2 will examine how PD genetic risk factors reported in the literature intersect with aSyn pathobiology, by testing the effect of knock-down or knock- in of individual genes on the formation of seeded pathology and neuronal survival. Candidates that significantly alter either will be confirmed in vivo using knock-out/knock-in mouse lines. Lastly, Aim 3 will integrate our molecular, genetic, and in vivo experimental data together with publicly available connectivity and gene- expression atlases to interrogate the mechanisms of pathological spread. Using recently developed mathematical approaches to describe infectious agent spread, we will develop in silico models to understand aSyn pathology formation and spread. Completion of these studies should provide valuable insights into the potential mechanisms by which aSyn contribute to the progression of PD and related disorders. Increased understanding of the pathogenesis of this and related synucleinopathies should ultimately result in earlier detection and disease-modifying therapies for these currently incurable disorders.