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dc.contributor.advisorCook, Gregory
dc.contributor.advisorArcus, Vickery
dc.contributor.authorFrampton, Rebekah Aimee
dc.date.available2011-10-17T00:32:06Z
dc.date.copyright2011
dc.identifier.citationFrampton, R. A. (2011). Toxin-Antitoxin Systems of Mycobacterium smegmatis (Thesis, Doctor of Philosophy). University of Otago. Retrieved from http://hdl.handle.net/10523/1900en
dc.identifier.urihttp://hdl.handle.net/10523/1900
dc.description.abstractThe genome of Mycobacterium smegmatis contains three putative toxin-antitoxin (TA) systems. The vapBC system has been characterized, but the remaining two TA systems, belonging to the mazEF and phd/doc family have not been studied. In this study, we show that overexpression of the mazF toxin caused killing in wild type cells, which was prevented by co-expressing the mazE antitoxin. Overexpression of doc was bacteriostatic, which was reversed by co-expression of phd. The mazEF and phd/doc genes were constitutively expressed, transcribed as leaderless transcripts, and subject to autoregulation. Taken together, these data demonstrate that these systems are also bona fide TA modules. No clear biological role has been assigned to TA systems encoded on bacterial chromosomes. To investigate the role of these TA modules in M. smegmatis, a triple knockout strain (ΔmazEF Δphd/doc ΔvapBC; ΔTAtriple) was constructed (strain RF105). No phenotypic differences were observed between wild type and ΔTAtriple when subjected to various stressors (e.g. antibiotics, pH, and hypoxia). However, long-term survival (viability) experiments in complex medium revealed a survival defect in ΔTAtriple compared to wild type cells demonstrating an essential role for these TA modules in mycobacterial adaptation to the conditions encountered by non-growing cells in spent medium (e.g. high pH, low carbon, hypoxia). Strikingly, microarray analysis revealed no significant differences in gene expression between the wild type and ΔTAtriple under these conditions, but metabolomic analysis indicated several metabolic differences. These included variations in the intracellular levels of serine and glycine, amino acids implicated in adaptation of mycobacterial cells to altered redox potential (e.g. hypoxia, NADH/NAD+ ratio). These data support a role for TA modules in the regulation of metabolic flux in mycobacterial cells to achieve balanced redox homeostasis in complex medium where multiple carbon and energy sources are present. The metabolic targets of these TA modules remain to be elucidated.en_NZ
dc.language.isoenen_NZ
dc.publisherUniversity of Otago
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dc.subjectMycobacteriaen_NZ
dc.subjectToxin-Antitoxinen_NZ
dc.titleToxin-Antitoxin Systems of Mycobacterium smegmatisen_NZ
dc.typeThesis
dc.date.updated2011-10-16T22:59:10Z
thesis.degree.disciplineMicrobiology and Immunologyen_NZ
thesis.degree.nameDoctor of Philosophyen_NZ
thesis.degree.grantorUniversity of Otago
thesis.degree.levelDoctoral
otago.interloanno
otago.openaccessAbstract Only
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