Mycobacterium abscessus glycopeptidolipids inhibit macrophage apoptosis and bacterial spreading by targeting mitochondrial cyclophilin D

Chul Hee Choi,Kang-In Lee,Nagatoshi Fujiwara,Yong Woo Back,Hwa-Jung Kim,Seungwha Paik,Jake Whang,Jeong-Kyu Park

doi:10.1038/cddis.2017.420

Chul Hee Choi, Kang-In Lee + Show 6 more

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https://doi.org/10.1038/cddis.2017.420

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Abstract

Mycobacterium abscessus (MAB) is a species of nontuberculous mycobacteria (NTM) and a major causative pathogen of pulmonary diseases especially in patients with cystic fibrosis. MAB infection is notoriously difficult to treat because of its intrinsic or inducible resistance to most antibiotics. The rough (R) morphotype of MAB, lacking cell surface glycopeptidolipids (GPLs), is associated with more severe and persistent infection than the smooth (S) type; however, the mechanisms underlying the R type’s virulence and the relation with GPLs remain unclear. In this study, we found that R-type MAB is much more proapoptotic than the S type, as a result of GPL-mediated inhibition of macrophage apoptosis. Polar GPLs inhibited an apoptotic response (induced by proapoptotic stimuli) by suppressing ROS production and the cytochrome c release and by preserving mitochondrial transmembrane potential. Furthermore, GPLs were found to be targeted to mitochondria and interacted with cyclophilin D; their acetylation was essential for this interaction. Finally, GPLs inhibited the intracellular growth and bacterial spreading of R-type MAB among macrophages via apoptosis inhibition. These findings suggest that GPLs limit MAB virulence by inhibiting apoptosis and the spread of bacteria and therefore provide a novel insight into the mechanism underlying virulence of MAB.

Highlights

Mycobacterium abscessus (MAB) is a rapidly growing nontuberculous mycobacterium (NTM) and is a common causative agent of lung diseases especially in cystic fibrosis patients.[1,2] MAB infection is notoriously difficult to treat because of its intrinsic or inducible resistance to most antibiotics.[3,4] the isolation rate of NTM species such as M. avium (MAV), MAB, and M. kansasii in clinical samples is rapidly increasing, little is known about their virulence factors and their pathogenicity at the cellular level
RAW 264.7 cells were infected with MAB at multiplicity of infections (MOIs) of 1 and 5, and their cell death pattern was analyzed by fluorescenceactivated cell sorting (FACS) with annexin V/propidium iodide (PI) staining from 1 to 4 h with 1 h intervals
The difference in the cell death induced by R and S types was not due to a difference in bacterial clumping as indicated by acid fast bacilli (AFB) staining of both bacterial types prepared for infection (Supplementary Figure 1a), and there was no difference in the macrophage infection rate between the two types (Supplementary Figure 1b)

Summary

Introduction

Mycobacterium abscessus (MAB) is a rapidly growing nontuberculous mycobacterium (NTM) and is a common causative agent of lung diseases especially in cystic fibrosis patients.[1,2] MAB infection is notoriously difficult to treat because of its intrinsic or inducible resistance to most antibiotics.[3,4] the isolation rate of NTM species such as M. avium (MAV), MAB, and M. kansasii in clinical samples is rapidly increasing, little is known about their virulence factors and their pathogenicity at the cellular level. The GPL core forms apolar or nonspecific GPLs (nsGPLs) that are produced by all NTM species, whereas polar, serotype-specific GPLs (ssGPLs) vary depending on the strain.[12,13] MAB produces unique polar GPLs that are diglycosylated on alaninol and di-O-acetylated on 6-deoxytalose.[12,13] MAB GPLs mask underlying bioactive cell wall lipids, thereby delaying the immunity activation during the initial colonization stages,[14] indicating their role as a virulence factor. A study on a zebrafish infection model indicated that R-type MAB induces more macrophage apoptosis than the S type and the virulence correlates with a loss of GPL production and massive production of a serpentine cord.[22] In the present study, we examined the precise role of GPLs in virulence and why the R type is more virulent than S type. We used extracted GPLs, studied their targeting to mitochondria, and their function in macrophage apoptosis

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