PROJECT SUMMARY / ABSTRACT Tuberculosis (TB) is the leading infectious killer worldwide. Upon exposure to Mycobacterium tuberculosis (Mtb), most people develop asymptomatic latent TB infection (LTBI phenotype). However, Dr. Anterasian's collaborators identified about 7% of individuals who, despite household exposure to pulmonary TB, do not convert their tuberculin skin test (TST) or interferon-γ release assay (IGRA), and thus can be classified as clinically resistant to traditionally defined LTBI (“RSTR” phenotype). Uncovering mechanisms of natural resistance to Mtb infection may provide unique insights that can inform the development of host-directed therapeutics (HDTs). Dr. Anterasian has analyzed the first global proteomic dataset of Mtb-infected primary human macrophages and discovered 46 differentially abundant proteins (DAPs) that define the RSTR vs LTBI macrophage proteomic response to Mtb. By integrating her data with those of her collaborators, Dr. Anterasian has identified DAPs that may undergo Mtb-induced post-translational modifications (PTMs) and/or interact with Mtb bacterial proteins. She has also used bioinformatic network analyses as well as genetic and transcriptomic data from the same patient cohort to ultimately curate a list of 19 proteins for further mechanistic studies. In particular, the Rab family of GTPases interact with Mtb bacterial proteins, undergo Mtb-induced ubiquitination, and are key regulators of membrane trafficking and autophagy, which are pathways Mtb exploits during infection. The objective of this proposal is to define macrophage pathways and proteins that characterize the protective RSTR response. Dr. Anterasian hypothesizes that RSTR individuals promote macrophage Mtb clearance through DAP-mediated pathways modulated by DAP-Mtb protein interactions and differential DAP ubiquitination. In Aim 1, Dr. Anterasian will investigate Rab-dependent mechanisms of protection against Mtb in macrophages and how Mtb proteins subvert Rab function. In Aim 2, Dr. Anterasian will identify candidate proteins associated with control of Mtb infection, their PTMs, and ubiquitin-dependent mechanisms of resistance. By combining proteomic, bioinformatic, and cellular immunology approaches, Dr. Anterasian is well poised to identify key pathways in human macrophage resistance to Mtb that can be targeted with HDTs. Dr. Anterasian is a Pediatric Infectious Disease Fellow in the Division of Infectious Diseases at Seattle Children's Hospital and the University of Washington. She additionally proposes a comprehensive career development program that includes: 1) mentored training in proteomics and cellular immunology; 2) formal didactics in large data set analyses (i.e. proteomics, statistical genetics, systems biology); 3) mentorship in the design of proteomic studies, human subjects research, and scientific writing; 4) structured opportunities to present her work to local and international scientific audiences; and 5) Scientific Advisory Committee meetings that ensure scientific and career development progress. By the conclusion of this award, Dr. Anterasian will transition to an independently-funded expert in the Mtb host response who will direct her own laboratory.

PROJECT NARRATIVE We have identified unique macrophage responses in humans who naturally resist Mycobacterium tuberculosis (Mtb) infection despite heavy exposure. The aims of this proposal are to uncover proteomic determinants of this macrophage resistance and define the underlying cellular mechanisms and Mtb-host protein interactions. Once key protective pathways are identified, they can be augmented with targeted host-directed therapies that improve killing of drug-sensitive and drug-resistant Mtb.