Proteomics and Network Analyses Reveal Inhibition of Akt‐mTOR Signaling in CD4+ T Cells by Mycobacterium tuberculosis Mannose‐Capped Lipoarabinomannan

• Published in: Proteomics (Weinheim) Vol. 17; no. 22
• Main Authors: Karim, Ahmad F., Sande, Obondo J., Tomechko, Sara E., Ding, Xuedong, Li, Ming, Maxwell, Sean, Ewing, Rob M., Harding, Clifford V., Rojas, Roxana E., Chance, Mark R., Boom, W. Henry
• Format: Journal Article
• Language: English
• Published: Germany Wiley Subscription Services, Inc 01-11-2017; John Wiley and Sons Inc
• Subjects: Akt; AKT protein; Animals; Bridges; CD28 antigen; CD3 antigen; CD4 antigen; CD4+ T‐cell; CD4-Positive T-Lymphocytes - metabolism; Cell activation; Cell Cycle; Cell walls; Costimulator; Female; Gene Regulatory Networks; Inhibition; label‐free mass spectrophotometry; Lipopolysaccharides - pharmacology; Lymphocytes; Lymphocytes T; M. tuberculosis; ManLAM; Mannose; Mannose - chemistry; Mass Spectrometry; Metabolism; Mice; Mice, Inbred C57BL; Modules; mTOR; Mycobacterium tuberculosis; Mycobacterium tuberculosis - metabolism; Network analysis; NF-κB protein; Oncogene Protein v-akt - antagonists & inhibitors; Oncogene Protein v-akt - metabolism; Peptides; Phosphorylation; Proliferating cell nuclear antigen; Proteins; Proteomics; Proteomics - methods; Ribonucleic acid; RNA; Signal Transduction; Signaling; Systems Biology; T cell receptors; TOR protein; TOR Serine-Threonine Kinases - antagonists & inhibitors; TOR Serine-Threonine Kinases - metabolism; Tricarboxylic acid cycle; Tuberculosis; mTOR; ManLAM; Akt; label-free mass spectrophotometry; CD4+ T-cell; M. tuberculosis
• ISSN: 1615-9853; 1615-9861
• DOI: 10.1002/pmic.201700233

Mycobacterium tuberculosis (Mtb) cell wall glycolipid mannose‐capped lipoarabinomannan (ManLAM) inhibits CD4+ T‐cell activation by inhibiting proximal T‐cell receptor (TCR) signaling when activated by anti‐CD3. To understand the impact of ManLAM on CD4+ T‐cell function when both the TCR–CD3 complex and major costimulator CD28 are engaged, we performed label‐free quantitative MS and network analysis. Mixed‐effect model analysis of peptide intensity identified 149 unique peptides representing 131 proteins that were differentially regulated by ManLAM in anti‐CD3‐ and anti‐CD28‐activated CD4+ T cells. Crosstalker, a novel network analysis tool identified dysregulated translation, TCA cycle, and RNA metabolism network modules. PCNA, Akt, mTOR, and UBC were found to be bridge node proteins connecting these modules of dysregulated proteins. Altered PCNA expression and cell cycle analysis showed arrest at the G2M phase. Western blot confirmed that ManLAM inhibited Akt and mTOR phosphorylation, and decreased expression of deubiquitinating enzymes Usp9x and Otub1. Decreased NF‐κB phosphorylation suggested interference with CD28 signaling through inhibition of the Usp9x‐Akt‐mTOR pathway. Thus, ManLAM induced global changes in the CD4+ T‐cell proteome by affecting Akt‐mTOR signaling, resulting in broad functional impairment of CD4+ T‐cell activation beyond inhibition of proximal TCR–CD3 signaling.