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Dynamic post-translational modification profiling of M. tuberculosis-infected primary macrophages

Citation

Budzik, Jonathan M et al. (2020), Dynamic post-translational modification profiling of M. tuberculosis-infected primary macrophages, Dryad, Dataset, https://doi.org/10.7272/Q6JQ0Z6J

Abstract

Macrophages are highly plastic cells with critical roles in immunity, cancer, and tissue homeostasis, but how these distinct cellular fates are triggered by environmental cues is poorly understood. To uncover how primary murine macrophages respond to bacterial pathogens, we globally assessed changes in post-translational modifications of proteins during infection with Mycobacterium tuberculosis, a notorious intracellular pathogen. We identified hundreds of dynamically regulated phosphorylation and ubiquitylation sites, indicating that dramatic remodeling of multiple host pathways, both expected and unexpected, occurred during infection. Most of these cellular changes were not captured by mRNA profiling, and included activation of ubiquitin-mediated autophagy, an evolutionarily ancient cellular antimicrobial system. This analysis also revealed that a particular autophagy receptor, TAX1BP1, mediates clearance of ubiquitylated Mtb and targets bacteria to LC3-positive phagophores. These studies provide a new resource for understanding how macrophages shape their proteome to meet the challenge of infection.

Methods

Immunofluorescence microscopy images were obtained with the Opera Phenix (Perkin Elmer) at 63X magnification.

Immunoblot images were obtained with a Li-Cor Odyssey instrument.

Usage Notes

Please see the manuscript file for details about staining protocols.

Funding

National Institute of Allergy and Infectious Diseases, Award: U19 AI106754

National Institute of Allergy and Infectious Diseases, Award: R01 AI120694

National Institute of Allergy and Infectious Diseases, Award: 1K08AI146267

National Institute of Allergy and Infectious Diseases, Award: DP1 AI124619

National Institute of Allergy and Infectious Diseases, Award: P01 AI063302

Cystic Fibrosis Foundation, Award: Harry Shwachman Award

National Institute of General Medical Sciences, Award: P50 GM082250