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Open Access Publications from the University of California

Scholarly Works (3 results)

  • Thesis
  • Peer Reviewed
UC Santa Cruz Electronic Theses and Dissertations (2019)

Iron is an essential element for most bacterial pathogens, including Yersinia. During infection, the mammalian immune system sequesters iron from invading pathogens; yet in response to this “nutritional immunity”, many pathogenic bacteria are able to colonize host tissue despite the decrease in iron availability. Additionally, when these pathogens travel from the outside environment into a mammalian host, they also encounter varying levels of oxygen depending on the tissue, or inflammation state. The aim of this dissertation is to improve our understanding of how bacterial pathogens couple iron and oxygen sending with virulence regulation. Specifically, I used the bacterial pathogen Yersinia pseudotuberculosis to determine how the type III secretion system (T3SS) is controlled in response to iron and oxygen. The T3SS is a bacterial injectisome apparatus used by human pathogenic Yersinia spp., to manipulate host defenses. Some of our recent studies suggest that Yersinia pseudotuberculosis uses the transcriptional regulator IscR to control the T3SS through direct regulation of the T3SS master regulator LcrF. IscR is a transcriptional regulator that binds different DNA sequence motifs, depending on its [2Fe-2S] cluster-coordinating (holo-) or clusterless (apo-) form. Iron availability and oxygen tension have been shown to affect E. coli IscR [2Fe-2S] cluster integrity, therefore impacting expression of IscR target genes4. Collectively, these data suggest that Y. pseudotuberculosis may use IscR to sense changes in oxygen tension and iron bioavailability to control critical virulence genes. However, it is not yet known whether changes in iron and oxygen availability alter expression or function of the T3SS or other virulence factors in Yersinia. Understanding how iron impacts microbial virulence can contribute to better guidelines for patient care in the context of anemia, iron supplementation, iron overload and inflammation.

The main goal of this dissertation is to determine the role of IscR on the regulation of the T3SS master regulator LcrF. The first part of my dissertation sought to study in vitro how both iron and oxygen availability affect the expression of iscR, lcrF and T3SS. We demonstrated that the presence of oxygen drives expression of iscR and this in turn upregulates lcrF expression and T3SS activity. Additionally, we showed that under anaerobic conditions, iscR expression is upregulated when iron is depleted, activating lcrF expression and, thus, T3SS expression. The second part of this dissertation was to determine if IscR control on T3SS is important for virulence in the mouse model of infection. To test this, we performed a bread feeding model of infection, which mimics oral intake of contaminated food. Our results suggest that regulation of the T3SS in response to iron and oxygen through IscR is critical for virulence.

    Cover page: How does Yersinia coordinate iron/oxygen sensing with virulence regulation?
    • Article
    • Peer Reviewed
    UC Santa Cruz Previously Published Works (2019)
    The enteropathogen Yersinia pseudotuberculosis and the related plague agent Y. pestis require the Ysc type III secretion system (T3SS) to subvert phagocyte defense mechanisms and cause disease. Yet type III secretion (T3S) in Yersinia induces growth arrest and innate immune recognition, necessitating tight regulation of the T3SS. Here we show that Y. pseudotuberculosis T3SS expression is kept low under anaerobic, iron-rich conditions, such as those found in the intestinal lumen where the Yersinia T3SS is not required for growth. In contrast, the Yersinia T3SS is expressed under aerobic or anaerobic, iron-poor conditions, such as those encountered by Yersinia once they cross the epithelial barrier and encounter phagocytic cells. We further show that the [2Fe-2S] containing transcription factor, IscR, mediates this oxygen and iron regulation of the T3SS by controlling transcription of the T3SS master regulator LcrF. IscR binds directly to the lcrF promoter and, importantly, a mutation that prevents this binding leads to decreased disseminated infection of Y. pseudotuberculosis but does not perturb intestinal colonization. Similar to E. coli, Y. pseudotuberculosis uses the Fe-S cluster occupancy of IscR as a readout of oxygen and iron conditions that impact cellular Fe-S cluster homeostasis. We propose that Y. pseudotuberculosis has coopted this system to sense entry into deeper tissues and induce T3S where it is required for virulence. The IscR binding site in the lcrF promoter is completely conserved between Y. pseudotuberculosis and Y. pestis. Deletion of iscR in Y. pestis leads to drastic disruption of T3S, suggesting that IscR control of the T3SS evolved before Y. pestis split from Y. pseudotuberculosis.
      Cover page: Iron availability and oxygen tension regulate the Yersinia Ysc type III secretion system to enable disseminated infection
      • Article
      • Peer Reviewed
      UC Santa Cruz Previously Published Works (2019)
      The enteropathogen Yersinia pseudotuberculosis and the related plague agent Y. pestis require the Ysc type III secretion system (T3SS) to subvert phagocyte defense mechanisms and cause disease. Yet type III secretion (T3S) in Yersinia induces growth arrest and innate immune recognition, necessitating tight regulation of the T3SS. Here we show that Y. pseudotuberculosis T3SS expression is kept low under anaerobic, iron-rich conditions, such as those found in the intestinal lumen where the Yersinia T3SS is not required for growth. In contrast, the Yersinia T3SS is expressed under aerobic or anaerobic, iron-poor conditions, such as those encountered by Yersinia once they cross the epithelial barrier and encounter phagocytic cells. We further show that the [2Fe-2S] containing transcription factor, IscR, mediates this oxygen and iron regulation of the T3SS by controlling transcription of the T3SS master regulator LcrF. IscR binds directly to the lcrF promoter and, importantly, a mutation that prevents this binding leads to decreased disseminated infection of Y. pseudotuberculosis but does not perturb intestinal colonization. Similar to E. coli, Y. pseudotuberculosis uses the Fe-S cluster occupancy of IscR as a readout of oxygen and iron conditions that impact cellular Fe-S cluster homeostasis. We propose that Y. pseudotuberculosis has coopted this system to sense entry into deeper tissues and induce T3S where it is required for virulence. The IscR binding site in the lcrF promoter is completely conserved between Y. pseudotuberculosis and Y. pestis. Deletion of iscR in Y. pestis leads to drastic disruption of T3S, suggesting that IscR control of the T3SS evolved before Y. pestis split from Y. pseudotuberculosis.
        Cover page: Iron availability and oxygen tension regulate the Yersinia Ysc type III secretion system to enable disseminated infection
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