The overall goal of the project is to understand why low complexity (LC) protein domains form amyloid-type fibrils. Several proteins with a LC domain aggregate in the neurons of patients with amyotrophic lateral sclerosis (ALS), causing cell death. The aggregates are composed of fibrils that look similar to the amyloid fibrils found Alzheimer's patients. However, the protein that aggregates in Alzheimer's is very different. FUS is one such protein involved in ALS. FUS has also been found to aggregate functionally, while amyloid proteins do not, which poses the following questions: What are the molecular similarities between FUS fibers and amyloid protein fibers. What differences allow FUS to have functional interactions with other proteins? Through this project, Dr. Murray will become an expert in the structural biology of proteins with a LC domain. This is an exciting new field, with great biomedical significance for ALS and mRNA processing. The postdoctoral training will allow Dr. Murray to independently lead an active research group focused on the molecular basis for disease and normal biological function. Through his training, he will become an expert in a variety of biophysical techniques, strengthen his teaching skills, broadly disseminate the results of his research, and form collaborations for his future career as a leader in the structural biolog of protein aggregation. The following specific aims will be addressed by Dr. Murray's research:
Aim #1 : Obtain an atomic resolution structure for the fibers formed by FUS. A series of nuclear magnetic resonance (NMR) experiments will be performed to measure structural restraints for the FUS fibrils. The structural restraints will then be used to calculate an atomic resolution structure for the FUS fibrils.
Aim #2 : Determine changes in FUS fibril structure upon interaction with the RNA polymerase. The RNA polymerase contains a LC domain that interacts with the FUS LC domain in vivo. NMR and electron microscopy experiments will be performed to identify the nature of the molecular interactions by which FUS interacts with the RNA polymerase.
Aim #3 : Observe changes in FUS fibril structure in the G159E ALS mutant. The ALS G159E FUS mutant forms fibrils with greater stability than the wild type protein. The G159E mutant will be structurally characterized using NMR and electron microscopy and its stability will be probed using a detergent based assay. The project is relevant to the mission of the NIGMS because it will help further understanding of the molecular interactions by which amyloid-type fibers form. Fibers of this type are found in Alzheimer's, ALS, and type 2 diabetes patients. Detailed understanding of how these fibers form is integral for treatment of these diseases. The training proposed here will allow Dr. Murray to actively pursue this goal as an independent investigator.
Low complexity protein domains are found in over 1/3 of human genes and over 60% of cancer related genes. Recently, these domains have been highlighted as important, functional, protein domains that can localize cell functions such as mRNA processing and cell stress response, without the need for a physical barrier like a membrane. A subclass of these protein domains have been discovered to form amyloid-type structures like those found in Alzheimer's and Parkinson's but, unlike traditional amyloid, these structures are reversible; this project will address the physical differences low complexity domains have with traditional amyloid and present a generally applicable method to investigate low complexity domain structure, stability, and function.