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In the Hadjifrangiskou Lab (or in short, the H-team), we are interested in understanding regulatory mechanisms that underlie multicellular behavior and virulence in bacteria that cause urinary tract infections (UTI).

UTIs are among the most frequent bacterial infections, are highly prevalent among women and have a high degree of recurrence. Currently, antibiotics are the primary treatment option for UTI, however they oftentimes fail to eliminate infection, they perturb the host microbial flora and select for increased antibiotic resistance. This means that there is a pressing need for the development of alternative strategies for preventing and/or treating UTIs. Uropathogenic Escherichia coli (UPEC), which cause the vast majority of community-acquired and catheter-associated UTIs, have evolved a remarkable mechanism to evade host immune defenses and establish infection, by forming biofilm-like intracellular bacterial communities inside bladder cells, in addition to forming extracellular biofilms on host cell surfaces and on catheter implants.

We are interested in identifying and dissecting the mechanisms by which UPEC can deftly sense and respond to multiple sensory inputs during infection that could serve as new drug targets. Specifically, we study the regulatory relationships that exist between two-component signaling networks, we investigate how these regulatory relationships stratify biofilm formation and we study how we can interfere with signaling cascades to reprogram bacteria and enhance targeting. To achieve these goals, we employ a combination of genetic, biochemical and mass spectrometric analyses with robust in vivo functional assays and three robust murine models of UTI.

Current Projects include:

(1) Characterization of the QseBC- PmrAB two-component system network and its role in pathogenesis

Two-component systems integrate signals, modulating gene expression changes to tailor adaptation to changing surrounding environments. In their simplest form, two-component systems consist of a sensor kinase that upon signal interception alters the phosphorylation state of a cognate response regulator, which then mediates transcription. Typically, two-component system interactions are restricted to cognate partners and non-cognate partner cross-talk while possible, is not strong. We have previously shown that in the case of QseBC, deletion of the QseC sensor in UPEC leads to gene dysregulation and virulence attenuation, due to constitutive signal-independent activation of the QseB response regulator by the PmrB sensor. We are currently exploring: A) The identification of QseC residues responsible for QseB deactivation. B) The basis of interaction between QseB and and PmrAB, and C) The role of the PmrAB-QseBC interactions in response to different and multiple sensory inputs.


(2) Dissection of surface-associated UPEC biofilms, using imaging mass spectrometry (IMS)

When bacteria form multicellular communities called biofilms they become less susceptible to host immune responses and antibiotic treatments. This recalcitrances stems in part from the protection afforded by a self-secreted extracellular matrix that shrouds the biofilm bacteria and retards penetration of the biomass by incoming stressors. Another factor that contributes to biofilm resilience is the presence of transient bacterial subpopulations that each provide the community with common goods and exhibit distinct characteristics. Identification of subpopulations and further characterization of their behavior will uncover potential biofilm markers, as well as targets for biofilm distortion/disassembly or inhibition. We are using IMS to detect areas with distinct protein profiles within preformed biofilms and we are using this information to understand how the immediate extracellular environment shapes the fate of these subpopulations within biofilms


(3) Investigation of previously uncharacterized UPEC biofilm effectors, the disruption of which diminishes IBC formation and attenuates UPEC virulence

Using a transposon screen, several UPEC biofilm determinants were discovered, which also impact UPEC IBC formation and in vivo virulence. We are interested in further characterizing each category of mutants and understanding the mechanism by which they modulate IBC formation.


(4) Understand the mechanism by which rationally designed small molecules act against UPEC and Helicobacter pylori transport systems

Our collaborator chemists have designed small molecular weight inhibitors to block UPEC pilus assembly, thereby impeding bacterial adherence to biotic and abiotic surfaces; we have come to discover that these molecules impact UPEC gene expression, causing pleiotropic effects on virulence gene regulation and inhibiting biofilm formation. In collaboration with labs at Washington University in Saint Louis and Umea University in Sweden, we are working towards understanding how these inhibitors function and how they can be optimized for use as treatments against UTIs. In parallel, we found that a subset of these compounds have profound effects on type 4 secretion systems in diverse extra-cellular pathogens.

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