TY - JOUR
T1 - Mycobacterium tuberculosis is protected from NADPH oxidase and LC3-associated phagocytosis by the LCP protein CpsA
AU - Köster, Stefan
AU - Upadhyay, Sandeep
AU - Chandra, Pallavi
AU - Papavinasasundaram, Kadamba
AU - Yang, Guozhe
AU - Hassan, Amir
AU - Grigsby, Steven J.
AU - Mittal, Ekansh
AU - Park, Heidi S.
AU - Jones, Victoria
AU - Hsu, Fong Fu
AU - Jackson, Mary
AU - Sassetti, Christopher M.
AU - Philips, Jennifer A.
N1 - Funding Information:
ACKNOWLEDGMENTS. We thank M. Dinauer (WUSM), H. W. Virgin (WUSM), D. Young (St. Jude’s Children’s Research Hospital), N. Mizushima (University of Tokyo), A. Yamamoto (Columbia University), and K. Cadwell (NYU SOM) for their generosity in providing mice; W. Jacobs, Jr. (Albert Einstein College of Medicine) for the ΔkatG strain; BEI Resources for the anti-KatG antibody; J. Ernst (NYU SOM) for the anti-Ag85 antibody; M. Dinauer, H. W. Virgin, L. D. Sibley (WUSM), A. C. M. Boon (WUSM), and members of the J.A.P. laboratory for useful discussions and helpful comments on the manuscript; Jessica Chapman-Lim and Beatrix Ueberheide (NYU Proteomics Laboratory) for assistance with mass spectrometry; and Eric Tycksen and the Genome Technology Access Center (GTAC) in the Department of Genetics at WUSM for help with genomic analysis and violin plots. The work was supported by funding from the Potts Memorial Foundation (S.K.), Stony Wold-Herbert Fund (S.K.), NIH/NIAID Grant AI119670 (to M.J.), NIH/NIAID Grants AI107774 and AI064282 (to C.M.S.), NIH/NIAID Grant AI130454 (to J.A.P.), and by NYU SOM H37Rv or ΔcpsA. Lungs were harvested on day 17 (E), 20 (G and I), 21 (H), and 24 (F). In E and I, C57BL/6 (WT) mice were used as controls. n = 5 mice per group; data show mean ± SEM; #P < 0.10, *P ≤ 0.05, **P ≤ 0.01; ns, not significant; Mann–Whitney test.
Funding Information:
and WUSM. The protein mass spectrometry experiments were supported by NIH Shared Instrumentation Grant 1S10OD010582. Development of mice used in this publication was supported by NIH/NIAID Center of Excellence in Translational Research Award Number U19AI109725. The GTAC is partially sup-
Funding Information:
ported by National Cancer Institute Cancer Center Support Grant P30 CA91842, Institute of Clinical and Translational Sciences/Clinical and Translational Science Award Grant UL1TR000448 from the National Center for Research Resources, and NIH Roadmap for Medical Research.
Publisher Copyright:
© 2017, National Academy of Sciences. All rights reserved.
PY - 2017/10/10
Y1 - 2017/10/10
N2 - Mycobacterium tuberculosis’ success as a pathogen comes from its ability to evade degradation by macrophages. Normally macrophages clear microorganisms that activate pathogen-recognition receptors (PRRs) through a lysosomal-trafficking pathway called “LC3-associated phagocytosis” (LAP). Although M. tuberculosis activates numerous PRRs, for reasons that are poorly understood LAP does not substantially contribute to M. tuberculosis control. LAP depends upon reactive oxygen species (ROS) generated by NADPH oxidase, but M. tuberculosis fails to generate a robust oxidative response. Here, we show that CpsA, a LytR-CpsA-Psr (LCP) domain-containing protein, is required for M. tuberculosis to evade killing by NADPH oxidase and LAP. Unlike phagosomes containing wild-type bacilli, phagosomes containing the ΔcpsA mutant recruited NADPH oxidase, produced ROS, associated with LC3, and matured into antibacterial lysosomes. Moreover, CpsA was sufficient to impair NADPH oxidase recruitment to fungal particles that are normally cleared by LAP. Intracellular survival of the ΔcpsA mutant was largely restored in macrophages missing LAP components (Nox2, Rubicon, Beclin, Atg5, Atg7, or Atg16L1) but not in macrophages defective in a related, canonical autophagy pathway (Atg14, Ulk1, or cGAS). The ΔcpsA mutant was highly impaired in vivo, and its growth was partially restored in mice deficient in NADPH oxidase, Atg5, or Atg7, demonstrating that CpsA makes a significant contribution to the resistance of M. tuberculosis to NADPH oxidase and LC3 trafficking in vivo. Overall, our findings reveal an essential role of CpsA in innate immune evasion and suggest that LCP proteins have functions beyond their previously known role in cell-wall metabolism.
AB - Mycobacterium tuberculosis’ success as a pathogen comes from its ability to evade degradation by macrophages. Normally macrophages clear microorganisms that activate pathogen-recognition receptors (PRRs) through a lysosomal-trafficking pathway called “LC3-associated phagocytosis” (LAP). Although M. tuberculosis activates numerous PRRs, for reasons that are poorly understood LAP does not substantially contribute to M. tuberculosis control. LAP depends upon reactive oxygen species (ROS) generated by NADPH oxidase, but M. tuberculosis fails to generate a robust oxidative response. Here, we show that CpsA, a LytR-CpsA-Psr (LCP) domain-containing protein, is required for M. tuberculosis to evade killing by NADPH oxidase and LAP. Unlike phagosomes containing wild-type bacilli, phagosomes containing the ΔcpsA mutant recruited NADPH oxidase, produced ROS, associated with LC3, and matured into antibacterial lysosomes. Moreover, CpsA was sufficient to impair NADPH oxidase recruitment to fungal particles that are normally cleared by LAP. Intracellular survival of the ΔcpsA mutant was largely restored in macrophages missing LAP components (Nox2, Rubicon, Beclin, Atg5, Atg7, or Atg16L1) but not in macrophages defective in a related, canonical autophagy pathway (Atg14, Ulk1, or cGAS). The ΔcpsA mutant was highly impaired in vivo, and its growth was partially restored in mice deficient in NADPH oxidase, Atg5, or Atg7, demonstrating that CpsA makes a significant contribution to the resistance of M. tuberculosis to NADPH oxidase and LC3 trafficking in vivo. Overall, our findings reveal an essential role of CpsA in innate immune evasion and suggest that LCP proteins have functions beyond their previously known role in cell-wall metabolism.
KW - Autophagy
KW - LC3-associated phagocytosis
KW - LytR-CpsA-Psr
KW - M. tuberculosis
KW - NADPH oxidase
UR - http://www.scopus.com/inward/record.url?scp=85030792037&partnerID=8YFLogxK
U2 - 10.1073/pnas.1707792114
DO - 10.1073/pnas.1707792114
M3 - Article
C2 - 28973896
AN - SCOPUS:85030792037
SN - 0027-8424
VL - 114
SP - E8711-E8720
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 41
ER -