The
            <i>Mycobacterium marinum</i>
            ESX-1 system mediates phagosomal permeabilization and type I interferon production via separable mechanisms

Lienard, Julia; Nobs, Esther; Lovins, Victoria; Movert, Elin; Valfridsson, Christine; Carlsson, Fredric

doi:10.1073/pnas.1911646117

Cited by 60 publications

(93 citation statements)

References 52 publications

(92 reference statements)

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“…To the best of our knowledge, it is for the first time that EsxA acidic pH-dependent membrane insertion is observed inside the host cells, which is consistent to the results obtained in liposomes (28,29,42,48). Emerging evidence have demonstrated that EsxAB stays on the surface of mycobacteria and contributes to the virulence (26,27,49,50). Besides, on epithelial cells, endocytosis is usually related to bacteria's invasion (21,22).…”

Section: Discussionsupporting

confidence: 87%

Mycobacterial surface-associated ESX-1 virulence factors play a role in mycobacterial adherence and invasion into lung epithelial cells

Bao

Zhang

Wang

et al. 2020

Preprint

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EsxA has been recognized as an important virulence factor of Mycobacterium tuberculosis (Mtb) that plays an essential role in Mtb cytosolic translocation by penetrating phagosomal membranes with its acidic pH-dependent membrane permeabilizing activity (MPA). Currently, the reported cytolytic activity of EsxA at neutral pH is controversial. In this study we have obtained direct evidence that it is the ASB-14, a detergent used in EsxA purification, but not EsxA that causes cytolysis at neutral pH. We have also found that the exogenously added EsxA was internalized into lung epithelial cells (WI-26) and inserted into the host membranes, and these processes could be blocked by cytochalasin D and bafilomycin A. This indicates that EsxA is bound by host surface receptors and internalized into acidic endosomal compartments. This observation has intrigued us to investigate the role of EsxA in mycobacterial adherence and invasion in host cells. Interestingly, compared to the Mycobacterium marinum (Mm) wild type strain, the Mm strain with deletion of the esxBA operon (MmEsxA:B) had a lower adherence but a higher invasion in WI-26 cells. More interestingly, either inducible knockdown of EsxAB or removal of the bacterial surface-associated EsxAB by Tween-80 exhibited opposite results compared to gene knockout. Finally, the surface-associated EsxA is correlated to mycobacterial virulence. Together, the present study has shown for the first time that EsxA is internalized into the host cells and inserts into the host membranes, and mycobacterial surface-associated EsxAB plays an important role in mycobacterial adherence and invasion in host cells, which warrants further investigation.

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Section: Discussionsupporting

confidence: 87%

Mycobacterial surface-associated ESX-1 virulence factors play a role in mycobacterial adherence and invasion into lung epithelial cells

Bao

Zhang

Wang

et al. 2020

Preprint

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“…al. showed that blocking phagosomal acidification by Bafilomycin did not decrease the ESX-1-mediated phagosomal permeabilization, suggesting that acidification is not required for membrane permeabilization (39). Recently, Lienard et.…”

Section: Discussionmentioning

confidence: 99%

Nα-Acetylation of the virulence factor EsxA is required for mycobacterial cytosolic translocation and virulence

Aguilera

Karki

et al. 2020

Journal of Biological Chemistry

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The Mycobacterium tuberculosis virulence factor EsxA and its chaperone EsxB are secreted as a heterodimer (EsxA:B) and are crucial for mycobacterial escape from phagosomes and cytosolic translocation. Current findings support the idea that for EsxA to interact with host membranes, EsxA must dissociate from EsxB at low pH. However, the molecular mechanism by which the EsxA:B heterodimer separates is not clear. In the present study, using liposome-leakage and cytotoxicity assays, LC-MS/MS–based proteomics, and CCF-4 FRET analysis, we obtained evidence that the Nα-acetylation of the Thr-2 residue on EsxA, a post-translational modification that is present in mycobacteria but absent in Escherichia coli, is required for the EsxA:B separation. Substitutions at Thr-2 that precluded Nα-acetylation inhibited the heterodimer separation and hence prevented EsxA from interacting with the host membrane, resulting in attenuated mycobacterial cytosolic translocation and virulence. Molecular dynamics simulations revealed that at low pH, the Nα-acetylated Thr-2 makes direct and frequent “bind-and-release” contacts with EsxB, which generates a force that pulls EsxB away from EsxA. In summary, our findings provide evidence that the Nα-acetylation at Thr-2 of EsxA facilitates dissociation of the EsxA:B heterodimer required for EsxA membrane permeabilization and mycobacterial cytosolic translocation and virulence.

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“…Within the macrophage, ESX-1 mediates permeabilization of the mycobacterial phagosome [16,17]. Permeabilization induces the cGAS/STING and AIM2/NLRP3 cytosolic signaling pathways, which promote production of cytokines that may enhance bacterial survival [18][19][20][21][22].…”

Section: Introductionmentioning

confidence: 99%

Mycobacterium marinum phthiocerol dimycocerosates enhance macrophage phagosomal permeabilization and membrane damage

et al. 2020

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Phthiocerol dimycocerosates (PDIMs) are a class of mycobacterial lipids that promote virulence in Mycobacterium tuberculosis and Mycobacterium marinum. It has recently been shown that PDIMs work in concert with the M. tuberculosis Type VII secretion system ESX-1 to permeabilize the phagosomal membranes of infected macrophages. As the zebrafish-M. marinum model of infection has revealed the critical role of PDIM at the host-pathogen interface, we set to determine if PDIMs contributed to phagosomal permeabilization in M. marinum. Using an ΔmmpL7 mutant defective in PDIM transport, we find the PDIM-ESX-1 interaction to be conserved in an M. marinum macrophage infection model. However, we find PDIM and ESX-1 mutants differ in their degree of defect, with the PDIM mutant retaining more membrane damaging activity. Using an in vitro hemolysis assay-a common surrogate for cytolytic activity, we find that PDIM and ESX-1 differ in their contributions: the ESX-1 mutant loses hemolytic activity while PDIM retains it. Our observations confirm the involvement of PDIMs in phagosomal permeabilization in M. marinum infection and suggest that PDIM enhances the membrane disrupting activity of pathogenic mycobacteria and indicates that the role they play in damaging phagosomal and red blood cell membranes may differ.

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The Mycobacterium marinum ESX-1 system mediates phagosomal permeabilization and type I interferon production via separable mechanisms

Cited by 60 publications

References 52 publications

Mycobacterial surface-associated ESX-1 virulence factors play a role in mycobacterial adherence and invasion into lung epithelial cells

Mycobacterial surface-associated ESX-1 virulence factors play a role in mycobacterial adherence and invasion into lung epithelial cells

Nα-Acetylation of the virulence factor EsxA is required for mycobacterial cytosolic translocation and virulence

Mycobacterium marinum phthiocerol dimycocerosates enhance macrophage phagosomal permeabilization and membrane damage

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