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Open Access Publications from the University of California

Scholarly Works (4 results)

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UCLA Electronic Theses and Dissertations (2017)

Burkholderia pseudomallei and Burkholderia thailandensis are related facultative intracellular pathogens that infect a variety of cell types, escape into the cytoplasm of host cells, and spread from cell-to-cell via type 6 secretion system (T6SS)-mediated cell fusion. Recent work has demonstrated a new mechanism of rapid intracellular motility dependent on a cryptic flagellar locus, fla2. In the first section of this dissertation, I show that the Fla2 system is present and expressed in Australian strains of B. pseudomallei and B. thailandensis. The results demonstrated that the fla2 locus encodes lateral flagella expressed intracellularly. This Fla2 system promotes swarming on semisolid agar. Lateral flagella encoded by the fla2 locus may provide a growth advantage for Burkholderia in environmental conditions, as wild-type B. thailandensis outcompeted a nonmotile ∆fliC2 derivative in a semisolid agar swarming competition assay.

In the next section of this dissertation I tested the effect of intracellular expression of lateral flagella on innate immune detection of Burkholderia. Innate immune mechanisms, particularly NLRC4-driven pyroptosis and production of IL-1β, are critical for the host response to B. pseudomallei infection. Such innate immune responses were previously attributed to type 3 secretion system (T3SS) stimuli deployed intracellularly. Here, I show that this response is at least partially dependent on fliC2, the flagellin gene encoded in the fla2 cluster in B. thailandensis and Australian strains of B. pseudomallei. While deletion of fliC2 in B. thailandensis had no measurable effect on pyroptosis or intracellular bacterial growth, it led to a nearly 33% decrease in IL-1β production in wild-type (wt) murine macrophages, without affecting IL-1β release in nlrc4-deficient macrophages. FliC2 did not appear to contribute similarly to IL-1β secretion during B. pseudomallei NAU14B-6 infection despite the immunogenicity of B. pseudomallei FliC2 ectopically expressed in a B. thailandensis ∆fliC2 strain. By quantitative RT-PCR, I demonstrate that intracellular expression of fliC2 is lower in the B. pseudomallei strains tested than in B. thailandensis. This difference may provide an explanation for the discrepancy between the innate immune response observed against FliC2 in B. thailandensis versus B. pseudomallei, leaving open the possibility that differences in fliC2 expression levels between B. pseudomallei strains may lead to strain-specific innate immune responses.

In the final section of this dissertation, I describe the role of a novel fla2 regulator on the global regulatory cascade involved in the B. pseudomallei intracellular life cycle. Overexpression of the flagellar response regulator frr activates the expression of lateral flagella, but decreases plaquing efficiency in both Fla2+ and Fla2- strains of B. pseudomallei and B. thailandensis. I show through quantitative RT-PCR that frr affects not only expression of lateral flagella components, but also polar flagella components from the Fla1 system utilized for extracellular swimming by Burkholderia and type 3 secretion system components required for escape from an initial endosome and from autophagosomes. These results suggest that frr plays a role as a master regulator controlling multiple aspects of Burkholderia virulence. In this chapter, I also investigate the role of other B. pseudomallei master regulators on subsets of virulence genes and demonstrate that each of these regulators exhibits crosstalk with other regulatory systems involved in the intracellular life cycle.

Together, the results described in this dissertation define and characterize a novel intracellular motility system in B. pseudomallei. The expression of lateral flagella by Burkholderia in the host cytoplasm is a unique and surprising feature for an intracellular pathogen, given the cytosolic surveillance mechanisms evolved in mammalian hosts to respond to flagella. In the concluding remarks of this dissertation, I discuss the status of B. pseudomallei as an “accidental” pathogen and how mechanisms evolved to survive and persist in specific ecological niches can contribute to or stand in the way of mammalian pathogenesis.

    Cover page: Characterization of an Intracellular Flagellar System in Pathogenic Burkholderia Species
    • Article
    • Peer Reviewed
    UC Irvine Previously Published Works (2024)
    At many research-intensive universities in North America, there is a disproportionate loss of minoritized undergraduate students from Science, Technology, Engineering, and Mathematics (STEM) majors. Efforts to confront this diversity, equity, and inclusion (DEI) challenge, such as faculty adoption of evidenced-based instructional approaches that promote student success, have been slow. Instructional and pedagogical change efforts at the academic department level have been demonstrated to be effective at enacting reform. One potential strategy is to embed change agent individuals within STEM departments that can drive change efforts. This study seeks to assess whether tenure-track, teaching-focused faculty housed in STEM departments are perceived as influential on the instructional and pedagogical domains of their colleagues. To answer this, individuals across five STEM departments at large, research-intensive campuses identified faculty who were influential upon six domains of their instruction and pedagogy. Social network analysis of individuals in these departments revealed heterogeneity across the instructional domains. Some, like the teaching strategies network, are highly connected and involve the majority of the department; while others, like the DEI influence network, comprise a significantly smaller population of faculty. Importantly, we demonstrate that tenure-track, teaching-focused faculty are influential across all domains of instruction, but are disproportionately so in the sparsely populated DEI influence networks.
      Cover page: Instructional Influencers: Teaching Professors as Potential Departmental Change Agents in Diversity, Equity, and Inclusion.
      • Article
      • Peer Reviewed
      UC San Diego Previously Published Works (2015)
      mRNA decapping is a central step in eukaryotic mRNA decay that simultaneously shuts down translation initiation and activates mRNA degradation. A major complex responsible for decapping consists of the decapping enzyme Dcp2 in association with decapping enhancers. An important question is how the activity and accumulation of Dcp2 are regulated at the cellular level to ensure the specificity and fidelity of the Dcp2 decapping complex. Here, we show that human Dcp2 levels and activity are controlled by a competition between decapping complex assembly and Dcp2 degradation. This is mediated by a regulatory domain in the Dcp2 C terminus, which, on the one hand, promotes Dcp2 activation via decapping complex formation mediated by the decapping enhancer Hedls and, on the other hand, targets Dcp2 for ubiquitin-mediated proteasomal degradation in the absence of Hedls association. This competition between Dcp2 activation and degradation restricts the accumulation and activity of uncomplexed Dcp2, which may be important for preventing uncontrolled decapping or for regulating Dcp2 levels and activity according to cellular needs.
        Cover page: Competition between Decapping Complex Formation and Ubiquitin-Mediated Proteasomal Degradation Controls Human Dcp2 Decapping Activity
        • Article
        • Peer Reviewed
        UC San Diego Previously Published Works (2020)
        Individuals who identify as lesbian, gay, bisexual, transgender, queer, and otherwise nonstraight and/or non-cisgender (LGBTQ+) have often not felt welcome or represented in the biology community. Additionally, biology can present unique challenges for LGBTQ+ students because of the relationship between certain biology topics and their LGBTQ+ identities. Currently, there is no centralized set of guidelines to make biology learning environments more inclusive for LGBTQ+ individuals. Rooted in prior literature and the collective expertise of the authors who identify as members and allies of the LGBTQ+ community, we present a set of actionable recommendations to help biologists, biology educators, and biology education researchers be more inclusive of individuals with LGBTQ+ identities. These recommendations are intended to increase awareness of LGBTQ+ identities and spark conversations about transforming biology learning spaces and the broader academic biology community to become more inclusive of LGBTQ+ individuals.
          Cover page: Fourteen Recommendations to Create a More Inclusive Environment for LGBTQ+ Individuals in Academic Biology
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