Limits...
A mycobacterium ESX-1-secreted virulence factor with unique requirements for export.

McLaughlin B, Chon JS, MacGurn JA, Carlsson F, Cheng TL, Cox JS, Brown EJ - PLoS Pathog. (2007)

Bottom Line: In contrast, disruption of Mh3866 or Mh3867 within the extRD1 locus prevents CFP-10 secretion without effect on EspBM.We also found interaction between Rv3879c and Rv3871, a component of the ESX-1 machine, suggesting a mechanism for the secretion of EspB.The results establish EspB as a substrate of ESX-1 that is required for virulence and growth in macrophages and suggests that the contribution of ESX-1 to virulence may arise from the secretion of multiple independent substrates.

View Article: PubMed Central - PubMed

Affiliation: Program in Microbial Pathogenesis and Host Defense, University of California San Francisco, San Francisco, California, United States of America.

ABSTRACT
Specialized secretion systems of pathogenic bacteria commonly transport multiple effectors that act in concert to control and exploit the host cell as a replication-permissive niche. Both the Mycobacterium marinum and the Mycobacterium tuberculosis genomes contain an extended region of difference 1 (extRD1) locus that encodes one such pathway, the early secretory antigenic target 6 (ESAT-6) system 1 (ESX-1) secretion apparatus. ESX-1 is required for virulence and for secretion of the proteins ESAT-6, culture filtrate protein 10 (CFP-10), and EspA. Here, we show that both Rv3881c and its M. marinum homolog, Mh3881c, are secreted proteins, and disruption of RD1 in either organism blocks secretion. We have renamed the Rv3881c/Mh3881c gene espB for ESX-1 substrate protein B. Secretion of M. marinum EspB (EspBM) requires both the Mh3879c and Mh3871 genes within RD1, while CFP-10 secretion is not affected by disruption of Mh3879c. In contrast, disruption of Mh3866 or Mh3867 within the extRD1 locus prevents CFP-10 secretion without effect on EspBM. Mutants that fail to secrete only EspBM or only CFP-10 are less attenuated in macrophages than mutants failing to secrete both substrates. EspBM physically interacts with Mh3879c; the M. tuberculosis homolog, EspBT, physically interacts with Rv3879c; and mutants of EspBM that fail to bind Mh3879c fail to be secreted. We also found interaction between Rv3879c and Rv3871, a component of the ESX-1 machine, suggesting a mechanism for the secretion of EspB. The results establish EspB as a substrate of ESX-1 that is required for virulence and growth in macrophages and suggests that the contribution of ESX-1 to virulence may arise from the secretion of multiple independent substrates.

Show MeSH

Related in: MedlinePlus

Growth of M. marinum Secretion Mutants in BMDMsBMDMs were infected with M. marinum strains as described in Materials and Methods at a multiplicity of infection of 1, and growth of bacteria was monitored over time as in Figure 1. Data are summarized from three independent experiments. Strains differed significantly by one-way ANOVA after 24 h, 48 h, 72 h, 96 h, and 120 h (p < 0.001 for each).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC1937011&req=5

ppat-0030105-g003: Growth of M. marinum Secretion Mutants in BMDMsBMDMs were infected with M. marinum strains as described in Materials and Methods at a multiplicity of infection of 1, and growth of bacteria was monitored over time as in Figure 1. Data are summarized from three independent experiments. Strains differed significantly by one-way ANOVA after 24 h, 48 h, 72 h, 96 h, and 120 h (p < 0.001 for each).

Mentions: Of the ten M. marinum extRD1 mutants we examined, MmΔRD1, espBM::tn, and Mh3871::tn were disrupted for the secretion of all three substrates: ESAT-6, CFP-10, and EspBM. In contrast, the Mh3879::tn mutant was disrupted only for the secretion of ESAT-6 and EspBM, while the Mh3866::tn and Mh3867::tn mutants were disrupted only for ESAT-6 and CFP-10 secretion. To assess the importance of the multiple ESX-1 substrates for growth in macrophages, we infected murine bone marrow–derived macrophages (BMDMs) with wild-type M. marinum, with strains lacking one secreted effector, or with strains lacking secretion of all the known ESX-1 substrates. As shown in Figure 3, MmΔRD1, espBM::tn, and Mh3871::tn, which fail to secrete all substrates, are more attenuated for growth in macrophages than Mh3866::tn, which still secretes EspBM, or Mh3879c::tn, which still secretes CFP-10. Therefore, we conclude that the various substrates of ESX-1 each contribute to virulence.


A mycobacterium ESX-1-secreted virulence factor with unique requirements for export.

McLaughlin B, Chon JS, MacGurn JA, Carlsson F, Cheng TL, Cox JS, Brown EJ - PLoS Pathog. (2007)

Growth of M. marinum Secretion Mutants in BMDMsBMDMs were infected with M. marinum strains as described in Materials and Methods at a multiplicity of infection of 1, and growth of bacteria was monitored over time as in Figure 1. Data are summarized from three independent experiments. Strains differed significantly by one-way ANOVA after 24 h, 48 h, 72 h, 96 h, and 120 h (p < 0.001 for each).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC1937011&req=5

ppat-0030105-g003: Growth of M. marinum Secretion Mutants in BMDMsBMDMs were infected with M. marinum strains as described in Materials and Methods at a multiplicity of infection of 1, and growth of bacteria was monitored over time as in Figure 1. Data are summarized from three independent experiments. Strains differed significantly by one-way ANOVA after 24 h, 48 h, 72 h, 96 h, and 120 h (p < 0.001 for each).
Mentions: Of the ten M. marinum extRD1 mutants we examined, MmΔRD1, espBM::tn, and Mh3871::tn were disrupted for the secretion of all three substrates: ESAT-6, CFP-10, and EspBM. In contrast, the Mh3879::tn mutant was disrupted only for the secretion of ESAT-6 and EspBM, while the Mh3866::tn and Mh3867::tn mutants were disrupted only for ESAT-6 and CFP-10 secretion. To assess the importance of the multiple ESX-1 substrates for growth in macrophages, we infected murine bone marrow–derived macrophages (BMDMs) with wild-type M. marinum, with strains lacking one secreted effector, or with strains lacking secretion of all the known ESX-1 substrates. As shown in Figure 3, MmΔRD1, espBM::tn, and Mh3871::tn, which fail to secrete all substrates, are more attenuated for growth in macrophages than Mh3866::tn, which still secretes EspBM, or Mh3879c::tn, which still secretes CFP-10. Therefore, we conclude that the various substrates of ESX-1 each contribute to virulence.

Bottom Line: In contrast, disruption of Mh3866 or Mh3867 within the extRD1 locus prevents CFP-10 secretion without effect on EspBM.We also found interaction between Rv3879c and Rv3871, a component of the ESX-1 machine, suggesting a mechanism for the secretion of EspB.The results establish EspB as a substrate of ESX-1 that is required for virulence and growth in macrophages and suggests that the contribution of ESX-1 to virulence may arise from the secretion of multiple independent substrates.

View Article: PubMed Central - PubMed

Affiliation: Program in Microbial Pathogenesis and Host Defense, University of California San Francisco, San Francisco, California, United States of America.

ABSTRACT
Specialized secretion systems of pathogenic bacteria commonly transport multiple effectors that act in concert to control and exploit the host cell as a replication-permissive niche. Both the Mycobacterium marinum and the Mycobacterium tuberculosis genomes contain an extended region of difference 1 (extRD1) locus that encodes one such pathway, the early secretory antigenic target 6 (ESAT-6) system 1 (ESX-1) secretion apparatus. ESX-1 is required for virulence and for secretion of the proteins ESAT-6, culture filtrate protein 10 (CFP-10), and EspA. Here, we show that both Rv3881c and its M. marinum homolog, Mh3881c, are secreted proteins, and disruption of RD1 in either organism blocks secretion. We have renamed the Rv3881c/Mh3881c gene espB for ESX-1 substrate protein B. Secretion of M. marinum EspB (EspBM) requires both the Mh3879c and Mh3871 genes within RD1, while CFP-10 secretion is not affected by disruption of Mh3879c. In contrast, disruption of Mh3866 or Mh3867 within the extRD1 locus prevents CFP-10 secretion without effect on EspBM. Mutants that fail to secrete only EspBM or only CFP-10 are less attenuated in macrophages than mutants failing to secrete both substrates. EspBM physically interacts with Mh3879c; the M. tuberculosis homolog, EspBT, physically interacts with Rv3879c; and mutants of EspBM that fail to bind Mh3879c fail to be secreted. We also found interaction between Rv3879c and Rv3871, a component of the ESX-1 machine, suggesting a mechanism for the secretion of EspB. The results establish EspB as a substrate of ESX-1 that is required for virulence and growth in macrophages and suggests that the contribution of ESX-1 to virulence may arise from the secretion of multiple independent substrates.

Show MeSH
Related in: MedlinePlus