title: Analysis of SecA2-dependent substrates in Mycobacterium marinum identifies protein kinase G (PknG) as a virulence effector.
abstract: 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 marinum secA2 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.
authors.affiliation: Department of Medical Microbiology and Infection Control, VU University Medical Center, van der Boechorststraat 7, 1081 BT, Amsterdam, The Netherlands; Department of Molecular Microbiology, Institute of Molecular Cell Biology, VU University, de Boelelaan 1085, 1081 HV, Amsterdam, The Netherlands.
authors.forename: Aniek D
authors.initials: AD
authors.lastname: van der Woude
authors.forename: Esther J M
authors.initials: EJ
authors.lastname: Stoop
authors.forename: Michael
authors.initials: M
authors.lastname: Stiess
authors.forename: Sen
authors.initials: S
authors.lastname: Wang
authors.forename: Roy
authors.initials: R
authors.lastname: Ummels
authors.forename: Gunny
authors.initials: G
authors.lastname: van Stempvoort
authors.forename: Sander R
authors.initials: SR
authors.lastname: Piersma
authors.forename: Alessandro
authors.initials: A
authors.lastname: Cascioferro
authors.forename: Connie R
authors.initials: CR
authors.lastname: Jiménez
authors.forename: Edith N G
authors.initials: EN
authors.lastname: Houben
authors.forename: Joen
authors.initials: J
authors.lastname: Luirink
authors.forename: Jean
authors.initials: J
authors.lastname: Pieters
authors.forename: Astrid M
authors.initials: AM
authors.lastname: van der Sar
authors.forename: Wilbert
authors.initials: W
authors.lastname: Bitter
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism*; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism*; D004251:DNA Transposable Elements; D004195:Disease Models, Animal; D055786:Gene Knockout Techniques; D015151:Immunoblotting; D026901:Membrane Transport Proteins / Q000378:metabolism*; D016254:Mutagenesis, Insertional; D009164:Mycobacterium Infections / Q000382:microbiology; D019910:Mycobacterium marinum / Q000378:metabolism* / Q000472:pathogenicity; D013379:Substrate Specificity; D037521:Virulence Factors / Q000378:metabolism*; D015027:Zebrafish
chemical_list: D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D026901:Membrane Transport Proteins; D037521:Virulence Factors; D017869:Cyclic GMP-Dependent Protein Kinases; D000251:Adenosine Triphosphatases; C442123:SecA2 protein, Mycobacterium