title: The resuscitation-promoting factors of Mycobacterium tuberculosis are required for virulence and resuscitation from dormancy but are collectively dispensable for growth in vitro.
abstract: Mycobacterium tuberculosis contains five resuscitation-promoting factor (Rpf)-like proteins, RpfA-E, that are implicated in resuscitation of this organism from dormancy via a mechanism involving hydrolysis of the peptidoglycan by Rpfs and partnering proteins. In this study, the rpfA-E genes were shown to be collectively dispensable for growth of M. tuberculosis in broth culture. The defect in resuscitation of multiple mutants from a 'non-culturable' state induced by starvation under anoxia was reversed by genetic complementation or addition of culture filtrate from wild-type organisms confirming that the phenotype was associated with rpf-like gene loss and that the 'non-culturable' cells of the mutant strains were viable. Other phenotypes uncovered by sequential deletion mutagenesis revealed a functional differentiation within this protein family. The quintuple mutant and its parent that retained only rpfD displayed delayed colony formation and hypersensitivity to detergent, effects not observed for mutants retaining only rpfE or rpfB. Furthermore, mutants retaining rpfD or rpfE were highly attenuated for growth in mice with the latter persisting better than the former in late-stage infection. In conjunction, these results are indicative of a hierarchy in terms of function and/or potency with the Rpf family, with RpfB and RpfE ranking above RpfD.
authors.affiliation: MRC/NHLS/WITS Molecular Mycobacteriology Research Unit, DST/NRF Centre of Excellence for Biomedical TB Research, School of Pathology, University of the Witwatersrand and the National Health Laboratory Service, Johannesburg 2000, South Africa.
authors.forename: Bavesh D
authors.initials: BD
authors.lastname: Kana
authors.forename: Bhavna G
authors.initials: BG
authors.lastname: Gordhan
authors.forename: Katrina J
authors.initials: KJ
authors.lastname: Downing
authors.forename: Nackmoon
authors.initials: N
authors.lastname: Sung
authors.forename: Galina
authors.initials: G
authors.lastname: Vostroktunova
authors.forename: Edith E
authors.initials: EE
authors.lastname: Machowski
authors.forename: Liana
authors.initials: L
authors.lastname: Tsenova
authors.forename: Michael
authors.initials: M
authors.lastname: Young
authors.forename: Arseny
authors.initials: A
authors.lastname: Kaprelyants
authors.forename: Gilla
authors.initials: G
authors.lastname: Kaplan
authors.forename: Valerie
authors.initials: V
authors.lastname: Mizrahi
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics / Q000502:physiology*; D015169:Colony Count, Microbial; D003470:Culture Media / Q000378:metabolism; D016207:Cytokines / Q000235:genetics / Q000502:physiology*; D003902:Detergents / Q000494:pharmacology; D005260:Female; D017353:Gene Deletion; D005816:Genetic Complementation Test; D006801:Humans; D008264:Macrophages / Q000382:microbiology; D051379:Mice; D050296:Microbial Viability / Q000235:genetics; D016254:Mutagenesis, Insertional; D009169:Mycobacterium tuberculosis / Q000187:drug effects / Q000235:genetics / Q000254:growth & development* / Q000472:pathogenicity*; D014376:Tuberculosis / Q000382:microbiology; D014774:Virulence; D037521:Virulence Factors / Q000235:genetics / Q000502:physiology*
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D003470:Culture Media; D016207:Cytokines; D003902:Detergents; D037521:Virulence Factors; C113704:resuscitation-promoting factor, bacteria