Research
Olga Morozova – Detection and structural analysis of misfolded tau protein in neurodegenerative diseases
Tau is a microtubule-associated protein that is localized in axons and is involved in neurite extension and maintenance. Tau misfolded into paired helical filaments (PHFs) is a pathological hallmark of more than 20 sporadic and familial neurodegenerative disorders including Alzheimer’s disease (AD) and frontotemporal dementia. Previous research shows that tau may be induced to misfold into distinct conformations with seeding. Here we explore how tau adopts multiple distinct fibril conformations maintained from nucleation to propagation. Preliminary work for this project includes the development of an assay for the sensitive detection and structural amplification of pathological tau in the brain and cerebrospinal fluid (CSF) of individuals with AD. We then characterized the fibril structure with multiple structural analysis techniques to understand the specific conformational changes in tau protein associated with this disease. With the initial evidence for some structural differences and a possibility of the propagation of seed structure in vitro, characterization of misfolded tau fibrils from other types of tauopathies will be implemented to gain better insight into the differences between the distinct disease pathologies that might lead to possible diagnostic techniques.
Kyle McHugh – Creating a Library of Transcription Factors to Explore Mechanisms of Cellular Reprogramming
and Neurogenesis
The human nervous system is composed of a multitude of different cell types that engage in a complicated network of inter-cellular signaling. Additionally, with recent advances in cellular reprogramming, ectopically expressed transcription factors may be used to induce terminally differentiated somatic cells to revert to a state of pluripotency similar to that of embryonic stem cells or trans-differentiate directly into another distinct cell type. Unfortunately, compared to the diversity of neural sub-types found in the brain, the knowledge of reprogramming cells into specific types of neurons is still somewhat limited. By generating a library of inducible transcription factors spanning a large range of tissue specificity and expression level, the phase space of reprogramming possibilities may be more thoroughly investigated, providing much new insight into the genetic regulatory mechanisms that determine cell type.
Kyle Doolan – Engineered Antibody Therapeutics for the Treatment of Prion Diseases
Cellular and protein engineering techniques provide a unique opportunity to investigate a mechanistic understanding of biological processes. Prion disease infectivity is one of the most unique biological processes in that it originates from a protein only pathogen in the absence of viral or cellular pathogens. In our work we seek to identify cellular and protein parameters which block the interaction of the pathogenic form of the prion protein (PrPSc) with the natively expressed prion protein (PrPC). Previous researchers have developed antibodies that bind PrPC and slow the conversion to PrPSc in cell culture and animal models, though few attempts have been made to engineer existing antibodies for greater therapeutic effect. Construction and evaluation of engineered antibodies to more effectively navigate the biological milieu and disrupt the PrPC- PrPSc protein-protein interaction will tie antibody parameters (eg. affinity and thermal stability) to biological function. This work will also enable the development of a quantitative mechanistic model of the kinetic and thermodynamic aspects of antibody based therapies. The model and methods developed here will likely apply more generally to other antibody therapies for more common neurological diseases such as Alzheimer’s and Parkinson’s disease.
Stefanie Berges – Development of a fluorescent bar-coding system for cell- based proteomic libraries
It is important to elucidate the complex and multifaceted molecular system that makes up the cell. Proteins constitute a significant portion of molecules in a cell, yet current proteomic tools fall short of those available for genomic analysis. Thus, new technologies for high-throughput, quantitative analysis of the proteome are critically needed. To this end, we will design and implement a fluorescent bar coding system for cell based proteomic libraries as an alternative to the current technology, which uses DNA technology to identify members of such libraries. Such a fluorescent bar coding system would allow one to identify all of the members of proteomic libraries which met given criteria in a single flow cytometry run, greatly improving the throughput and reducing the cost of library screening. Other benefits of designing a system which utilizes flow cytometry include the ability to directly couple the identification process to another fluorescent readout (e.g. protein expression level, cell health, calcium homeostasis, etc.) as well as the advantage of single cell analysis, which provides quantitative analysis of individual members of cell populations, rather than population averages.