- Genetic and spatial maps of alpha-synucleinopathy in distinct CNS cell types and genetic backgrounds. We have generated induced pluripotent stem cell (iPSc) lines from a number of patients with mutations at the alpha-synuclein locus, as well as numerous patients with sporadic synucleinopathies (including Parkinson’s disease and multiple system atrophy). We have optimized differentiation protocols for a number of distinct neuronal and glial cell types, and developed tractable iPSC models that rapidly develop aggregation (inclusion) pathologies. In this project, unbiased proteome-scale screening methods (including CRISPR-Cas9 genome editing and APEX-based in situ proteome labeling) will be optimized to identify genes that modify neurodegenerative phenotypes in these cells and proteins that interact with alpha-synuclein. The key question we will address is how the genetic and spatial interaction map of a misfolded protein is altered in distinct cell types.
- Cellular pathologies associated with distinct biophysical forms (“strains”) of alpha synuclein in stem cell-derived neurons and glia. Recently, considerable interest has arisen around the spread of alpha-syuclein from cell to cell in a “prionoid” fashion. It has even be proposed that distinct synucleinopathies arise from different biophysical forms, or “strains”, of misfolded alpha-synuclein. We are examining this hypothesis in a patient-derived model system. Genetic manipulation and single-cell longitudinal imaging are used to understand and track the consequences of alpha-synuclein aggregation.
- The effects of alpha-synuclein on mRNA processing and translation. A variety of model systems will be used, from in vitro systems to yeast to patient-derived neurons and glia to better understand how alpha synuclein’s interactions with mRNA binding proteins reflects its intrinsic biology and pathology.
- Neuroinflammatory signaling. Increasingly neuroinflammatory processes are considered to be critical in the progression of neurodegenerative diseases, but the specific ways in which neuroimmune signaling is engaged as a result of protein misfolding is unclear. We are investigating how alpha-synuclein may specifically engage pathologic glial and immune responses when it accumulates or misfolds.
- Functional genomics. In a systems biology approach, we cross-correlate molecular network data gleaned from our cellular models with human genomic data. Together with our collaborators, we develop biologically driven tools for genomic analysis and patient stratification. Since the misfolding of alpha-synuclein by definition is the central pathology of synucleinopathies, regardless of the cause, we take our molecular maps as a starting point for understand why specific individual succumb to these diseases (and others escape). This project utilizes genomic sequencing data from individual patients, multi-generation kindreds we follow with dominant alpha-synuclein mutations and larger scale genomic datasets. Our ultimate aim is to develop therapeutics that rationally target biologically defined subsets of patients.
We have generated iPSC from patients who harbor CAG tract (polyglutamine-encoding) expansion mutations that lead to neurodegenerative ataxias, including SCA-1, SCA-2, SCA-3, SCA-7, SCA-8 and DRPLA. Beginning with SCA-3 and DRPLA, we are interested in understanding the biological consequences of misfolding of the encoded proteins and the effects on the multi-protein complexes in which they below. Therapeutically, we are interested in whether genetic correction of these expansions is equivalent to knockdown, a project that has consequences for understanding the potential (and also the potential limitations) of antisense gene therapy for these diseases.