Genetic regulation of virulence in Francisella tularensis

dc.contributor.advisorKlose, Karl E.
dc.contributor.authorRodriguez, Stephen A.
dc.contributor.committeeMemberSeshu, Janakiram
dc.contributor.committeeMemberArulanandam, Bernard P.
dc.contributor.committeeMemberGuentzel, Neal
dc.contributor.committeeMemberDube, Peter H.
dc.descriptionThis item is available only to currently enrolled UTSA students, faculty or staff. To download, navigate to Log In in the top right-hand corner of this screen, then select Log in with my UTSA ID.
dc.description.abstract<italic>Francisella tularensis</italic> is the causal agent of the zoonotic disease tularemia, and a potential biological weapon. Because studying the molecular mechanisms of pathogenesis has been difficult due to a lack of techniques for targeted gene disruption, we have adapted a gene knockout system that efficiently targets genes for disruption in <italic>F. tularensis</italic>. This system utilizes mobile group II introns (targetrons) specifically optimized for <italic>F. tularensis</italic>. Results showed that this system is effective across <italic>F. tularensis</italic> subspecies, and enables simultaneous inactivation of duplicate genes, which should prove useful in targeting duplicated virulence genes in the highly virulent strains. The virulence of <italic>F. tularensis</italic> is dependent on the genes encoded within the <italic>Francisella</italic> Pathogenicity Island (FPI). The FPI has recently been shown to encode a secretion system related to Type VI secretion systems found in other pathogenic bacteria. A single FPI is found in <italic>F. tularensis</italic> spp. <italic>novicida</italic>, which has low virulence for humans, whereas two identical FPIs are found in the highly virulent <italic>F. tularensis</italic> spp. <italic>tularensis</italic>. We utilized FLP recombinase-mediated excision to remove the entire FPI from <italic>F. tularensis</italic> spp. <italic>novicida</italic>. The resultant &#8710;FPI mutant was defective for intramacrophage replication, and was attenuated for virulence in mice. These results emphasize the importance of the FPI to <italic>F. tularensis</italic> spp. <italic>novicida</italic> virulence. Moreover, the FLP recombinase-mediated excision technique should prove useful in analyzing the duplication of the FPI in <italic>F. tularensis</italic> spp. <italic>tularensis</italic>. The FPI gene <italic>iglE</italic> encodes a putative lipoprotein with a predicted lipobox that may be part of the T6SS-like secretion system. We constructed an <italic>iglE</italic> mutant <italic>F. tularensis</italic> spp. <italic>novicida</italic> strain, and demonstrated that <italic>iglE</italic> contributes to intramacrophage replication. Membrane fractionation of <italic>F. tularensis</italic> expressing IglE-FLAG demonstrated that this protein is found in the outer membrane (OM), but mutation of the putative site of lipidation (C22G) prevents OM localization of IglE-FLAG. Additionally, OM localization of another FPI protein, IglA, appeared to require IglE, suggesting that IglE may facilitate localization of IglA. Further experiments will clarify the contribution of iglE to the pathogenesis of <italic>F. tularensis</italic>. The genetic techniques that have been developed and the investigation of IglE should contribute to deciphering the virulence of <italic>F. tularensis</italic>.
dc.description.departmentIntegrative Biology
dc.format.extent169 pages
dc.subjectFrancisella tularensis
dc.subjectpathogenicity island
dc.subjectsecretion system
dc.subject.classificationMolecular biology
dc.titleGenetic regulation of virulence in Francisella tularensis
dcterms.accessRightspq_closed Biology of Texas at San Antonio of Philosophy


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