Analysis of SecA2-dependent substrates in Mycobacterium marinum identifies protein kinase G (PknG) as a virulence effector.

A.D. van der Woude, E.J.M. Stoop, M. Stiess, Q. Wang, R. Ummels, G. van Stempvoort, S.R. Piersma, A. Cascioferro, C.R. Jimenez, E.N.G. Houben, J. Luirink, J. Pieters, A.M. van der Sar, W. Bitter

Research output: Contribution to JournalArticleAcademicpeer-review

Abstract

Summary: The pathogenicity of mycobacteria is closely associated with their ability to export virulence factors. For this purpose, mycobacteria possess different protein secretion systems, including the accessory Sec translocation pathway, SecA2. Although this pathway is associated with intracellular survival and virulence, the SecA2-dependent effector proteins remain largely undefined. In this work, we studied a Mycobacterium marinumsecA2 mutant with an impaired capacity to initiate granuloma formation in zebrafish embryos. By comparing the proteomic profile of cell envelope fractions from the secA2 mutant with wild type M.marinum, we identified putative SecA2-dependent substrates. Immunoblotting procedures confirmed SecA2-dependent membrane localization for several of these proteins, including the virulence factor protein kinase G (PknG). Interestingly, phenotypical defects of the secA2 mutant are similar to those described for ΔpknG, including phagosomal maturation. Overexpression of PknG in the secA2 mutant restored its localization to the cell envelope. Importantly, PknG-overexpression also partially restored the virulence of the secA2 mutant, as indicated by enhanced infectivity in zebrafish embryos and restored inhibition of phagosomal maturation. These results suggest that SecA2-dependent membrane localization of PknG is an important determinant for M.marinum virulence. © 2013 John Wiley & Sons Ltd.
Original languageEnglish
Pages (from-to)280-295
JournalCellular Microbiology
Volume16
Issue number2
DOIs
Publication statusPublished - 2014

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Mycobacterium marinum
Cyclic GMP-Dependent Protein Kinases
Virulence
Mycobacterium
Virulence Factors
Zebrafish
Embryonic Structures
Granuloma
Immunoblotting
Proteomics
Membrane Proteins
Proteins
Membranes

Cite this

van der Woude, A.D. ; Stoop, E.J.M. ; Stiess, M. ; Wang, Q. ; Ummels, R. ; van Stempvoort, G. ; Piersma, S.R. ; Cascioferro, A. ; Jimenez, C.R. ; Houben, E.N.G. ; Luirink, J. ; Pieters, J. ; van der Sar, A.M. ; Bitter, W. / Analysis of SecA2-dependent substrates in Mycobacterium marinum identifies protein kinase G (PknG) as a virulence effector. In: Cellular Microbiology. 2014 ; Vol. 16, No. 2. pp. 280-295.
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title = "Analysis of SecA2-dependent substrates in Mycobacterium marinum identifies protein kinase G (PknG) as a virulence effector.",
abstract = "Summary: The pathogenicity of mycobacteria is closely associated with their ability to export virulence factors. For this purpose, mycobacteria possess different protein secretion systems, including the accessory Sec translocation pathway, SecA2. Although this pathway is associated with intracellular survival and virulence, the SecA2-dependent effector proteins remain largely undefined. In this work, we studied a Mycobacterium marinumsecA2 mutant with an impaired capacity to initiate granuloma formation in zebrafish embryos. By comparing the proteomic profile of cell envelope fractions from the secA2 mutant with wild type M.marinum, we identified putative SecA2-dependent substrates. Immunoblotting procedures confirmed SecA2-dependent membrane localization for several of these proteins, including the virulence factor protein kinase G (PknG). Interestingly, phenotypical defects of the secA2 mutant are similar to those described for ΔpknG, including phagosomal maturation. Overexpression of PknG in the secA2 mutant restored its localization to the cell envelope. Importantly, PknG-overexpression also partially restored the virulence of the secA2 mutant, as indicated by enhanced infectivity in zebrafish embryos and restored inhibition of phagosomal maturation. These results suggest that SecA2-dependent membrane localization of PknG is an important determinant for M.marinum virulence. {\circledC} 2013 John Wiley & Sons Ltd.",
author = "{van der Woude}, A.D. and E.J.M. Stoop and M. Stiess and Q. Wang and R. Ummels and {van Stempvoort}, G. and S.R. Piersma and A. Cascioferro and C.R. Jimenez and E.N.G. Houben and J. Luirink and J. Pieters and {van der Sar}, A.M. and W. Bitter",
year = "2014",
doi = "10.1111/cmi.12221",
language = "English",
volume = "16",
pages = "280--295",
journal = "Cellular Microbiology",
issn = "1462-5814",
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van der Woude, AD, Stoop, EJM, Stiess, M, Wang, Q, Ummels, R, van Stempvoort, G, Piersma, SR, Cascioferro, A, Jimenez, CR, Houben, ENG, Luirink, J, Pieters, J, van der Sar, AM & Bitter, W 2014, 'Analysis of SecA2-dependent substrates in Mycobacterium marinum identifies protein kinase G (PknG) as a virulence effector.' Cellular Microbiology, vol. 16, no. 2, pp. 280-295. https://doi.org/10.1111/cmi.12221

Analysis of SecA2-dependent substrates in Mycobacterium marinum identifies protein kinase G (PknG) as a virulence effector. / van der Woude, A.D.; Stoop, E.J.M.; Stiess, M.; Wang, Q.; Ummels, R.; van Stempvoort, G.; Piersma, S.R.; Cascioferro, A.; Jimenez, C.R.; Houben, E.N.G.; Luirink, J.; Pieters, J.; van der Sar, A.M.; Bitter, W.

In: Cellular Microbiology, Vol. 16, No. 2, 2014, p. 280-295.

Research output: Contribution to JournalArticleAcademicpeer-review

TY - JOUR

T1 - Analysis of SecA2-dependent substrates in Mycobacterium marinum identifies protein kinase G (PknG) as a virulence effector.

AU - van der Woude, A.D.

AU - Stoop, E.J.M.

AU - Stiess, M.

AU - Wang, Q.

AU - Ummels, R.

AU - van Stempvoort, G.

AU - Piersma, S.R.

AU - Cascioferro, A.

AU - Jimenez, C.R.

AU - Houben, E.N.G.

AU - Luirink, J.

AU - Pieters, J.

AU - van der Sar, A.M.

AU - Bitter, W.

PY - 2014

Y1 - 2014

N2 - Summary: The pathogenicity of mycobacteria is closely associated with their ability to export virulence factors. For this purpose, mycobacteria possess different protein secretion systems, including the accessory Sec translocation pathway, SecA2. Although this pathway is associated with intracellular survival and virulence, the SecA2-dependent effector proteins remain largely undefined. In this work, we studied a Mycobacterium marinumsecA2 mutant with an impaired capacity to initiate granuloma formation in zebrafish embryos. By comparing the proteomic profile of cell envelope fractions from the secA2 mutant with wild type M.marinum, we identified putative SecA2-dependent substrates. Immunoblotting procedures confirmed SecA2-dependent membrane localization for several of these proteins, including the virulence factor protein kinase G (PknG). Interestingly, phenotypical defects of the secA2 mutant are similar to those described for ΔpknG, including phagosomal maturation. Overexpression of PknG in the secA2 mutant restored its localization to the cell envelope. Importantly, PknG-overexpression also partially restored the virulence of the secA2 mutant, as indicated by enhanced infectivity in zebrafish embryos and restored inhibition of phagosomal maturation. These results suggest that SecA2-dependent membrane localization of PknG is an important determinant for M.marinum virulence. © 2013 John Wiley & Sons Ltd.

AB - Summary: The pathogenicity of mycobacteria is closely associated with their ability to export virulence factors. For this purpose, mycobacteria possess different protein secretion systems, including the accessory Sec translocation pathway, SecA2. Although this pathway is associated with intracellular survival and virulence, the SecA2-dependent effector proteins remain largely undefined. In this work, we studied a Mycobacterium marinumsecA2 mutant with an impaired capacity to initiate granuloma formation in zebrafish embryos. By comparing the proteomic profile of cell envelope fractions from the secA2 mutant with wild type M.marinum, we identified putative SecA2-dependent substrates. Immunoblotting procedures confirmed SecA2-dependent membrane localization for several of these proteins, including the virulence factor protein kinase G (PknG). Interestingly, phenotypical defects of the secA2 mutant are similar to those described for ΔpknG, including phagosomal maturation. Overexpression of PknG in the secA2 mutant restored its localization to the cell envelope. Importantly, PknG-overexpression also partially restored the virulence of the secA2 mutant, as indicated by enhanced infectivity in zebrafish embryos and restored inhibition of phagosomal maturation. These results suggest that SecA2-dependent membrane localization of PknG is an important determinant for M.marinum virulence. © 2013 John Wiley & Sons Ltd.

U2 - 10.1111/cmi.12221

DO - 10.1111/cmi.12221

M3 - Article

VL - 16

SP - 280

EP - 295

JO - Cellular Microbiology

JF - Cellular Microbiology

SN - 1462-5814

IS - 2

ER -