Abstract

Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, is the leading cause of death from an infectious agent. Mtb is a major global health threat, killing nearly 1.25 million individuals yearly. Although tuberculosis can be deadly and is highly infectious, 90% of infected individuals have the capacity to clear infection or live asymptomatically with latent TB infection (LTBI). However, a small subset of individuals, 5-10%, are unable to clear infection and go on to develop active TB disease. The success of Mtb as a pathogen stems in part from its ability to evade innate immune defenses and establish long-term infection, specifically within macrophages. Following uptake by phagocytic cells, the bacilli are enclosed within phagosomes that progressively mature through a series of membrane trafficking events. Ultimately, phagosomes fuse with lysosomes to form phagolysosomes, degradative compartments enriched with antimicrobial properties that promote microbial killing. Despite macrophage effector functions that typically restrict microbial replication, Mtb produces a variety of protein and lipid effectors that manipulate host trafficking pathways such as autophagy. By preventing phagosome maturation and limiting phagosome-lysosome fusion, the bacteria are able to evade degradation and survive intracellularly. Accumulating evidence points to the importance of Mtb undermining lysosomal trafficking pathways LC3-associated phagocytosis (LAP) and selective autophagy (xenophagy). LAP is a form of non-canonical autophagy that is dependent on the recruitment of the NADPH oxidase which generates ROS and contributes to bacterial killing. In contrast, xenophagy is a selective form of canonical autophagy that recognizes intracellular pathogens following phagosomal damage or cytosolic exposure. These pathogens are sequestered into autophagosomal membranes and subsequently delivered to lysosomes, where they are degraded. Previously, we demonstrated that the Mtb protein, CpsA, is critical for bacterial survival in macrophages and infection in mice, and that it can inhibit LAP by inhibiting the recruitment of the NADPH oxidase and ROS generation. CpsA belongs to the LCP family of proteins involved in cell envelope maintenance, but our work demonstrated that it has evolved to have a function beyond that of a cell wall ligase. In addition to inhibiting LAP, we also found that CpsA can interact with NDP52 and TAX1BP1, autophagy receptors that recognize cytosolic bacteria and target them for lysosomal degradation, raising the possibility that CpsA also disrupts canonical autophagy. Here, we show that CpsA binds NDP52 and TAX1BP1 in their SKICH domains, which normally recruit the autophagy initiation machinery and the TBK1 kinase which, are required for autophagosome formation. We demonstrate that CpsA prevents phagosomal recruitment of FIP200, a component of the autophagy initiation machinery, and blocks TBK1-dependent phosphorylation of p62 at serine 403, leading to impaired xenophagy. Our findings provide mechanistic insight into how Mtb subverts host defenses, revealing that in addition to blocking non-canonical autophagy, CpsA inhibits the TBK1-p62 axis to block xenophagy; thereby disarming both arms of lysosomal trafficking in macrophages.

Committee Chair

Jennifer Philips

Committee Members

Anthony Orvadahl; Christina Stallings; Daniel Goldberg; Jeffrey Henderson

Degree

Doctor of Philosophy (PhD)

Author's Department

Biology & Biomedical Sciences (Molecular Microbiology & Microbial Pathogenesis)

Author's School

Graduate School of Arts and Sciences

Document Type

Dissertation

Date of Award

8-11-2026

Language

English (en)

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Available for download on Wednesday, February 10, 2027

Included in

Microbiology Commons

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