Defining the role of site-specific proteolysis in innate defense signaling

PROJECT SUMMARY Human cells respond to foreign agents such as pathogens and toxins by initiating a strong innate defense response that creates a protective environment in the cells and incapacitates the invading pathogens and foreign substances. The initiation, activation, and resolution of this innate defense response is a carefully regulated process designed to avoid both hyperactivation and underactivation, either of which can lead to tissue damage, organ dysfunction, and microbial diseases. A key strategy that cells use to achieve tight regulatory control of innate defense signaling is the alteration of protein post-translational modifications (PTMs: phosphorylation, ubiquitination, acetylation, etc.). The goal of my lab for the next five years will be to explore the role of an understudied PTM, the site-specific proteolysis, in regulating innate defense signaling. I have extensive experience in studying antiviral innate immune responses, and have recently optimized a robust protein N-terminal labeling approach that provides an unbiased view of site-specific proteolysis in cells — expertise that will enable me to decipher the regulatory functions of site-specific proteolysis in innate defense mechanisms. With this MIRA award, my group will pursue two specific research directions. First, we will determine the role of the proteolytic N-end rule pathway in controlling the stability of proteins involved in defense signaling. Second, we will investigate the ways in which internal protein cleavages activate defense responses. For the proposed studies, we will employ a combination of focused molecular biology investigations and systems biology approaches, such as activity-based protease profiling and protein N-terminal labeling. These efforts will generate new knowledge about the role of proteases in the regulation of cellular defenses and inform the development of strategies to improve the performance of innate defense mechanisms against escalating microbial and environmental threats.