Graduate Students Mini Symposium V-2024

01:15 PM Camila Cilveti Rodriguez Riglos (PG Yuan)
The involvement of PhoPQ and MgrB in the colonization process of E. coli

Uropathogenic E. coli (UPEC) is one of the most common etiological agents of urinary tract
infections (UTIs). UTIs often begin with contamination of an uropathogen residing in the gut that
acts as a reservoir. However, the gut colonization of UPEC is not yet entirely characterized.
Bacteria primarily utilize two-component signaling pathways to adapt to the continuously changing
environment of the gastrointestinal tract. Among them is the PhoQ/PhoP two-component system,
which is involved in magnesium homeostasis, stress response, resistance to cationic antimicrobial
peptides, long-chain fatty acid response, and virulence regulation. The PhoQ/PhoP system
triggers a signaling cascade that results in gene expression, enabling bacteria to cope with
environmental threats and nutritional deficiencies. Using proteomic and molecular approaches,
we identified the regulation of F9 fimbriae operon by the response regulator PhoP. F9 fimbriae
are hair-like proteinous appendages on the cell surface, allowing bacteria to attach to specific
receptors in the host tissue, thus playing critical roles in the colonization process. Consistently,
using in vitro adhesion assays, we identified an increase in bacterial attachment to the
gastrointestinal epithelial barrier when the PhoQ/PhoP system was at or near a fully active state,
suggesting the involvement of PhoPQ and MgrB in the initial attachment in the gut. Through this
work, we aim to contribute to the knowledge about the PhoPQ regulatory network, uncover the
importance of PhoPQ negative regulator, MgrB, and their physiological effects in the colonization
of pathogenic and commensal E. coli.

01:45 PM Xuanlin Chen (AG Sourjik)

Pulsatile flagellar gene expression drives c-di-GMP dynamics in single bacterial cells

Escherichia coli employs c-di-GMP to modulate motile-to-sessile lifestyle switching. The classic motility/biofilm network represents a deterministic and mutually exclusive regulation of flagellar gene expression and c-di-GMP production. However, FliA (flagellar sigma factor) provides both negative and positive regulation into c-di-GMP via PdeH (the major phosphodiesterase) and RdcA (regulator of the major diguanylate cyclase, DgcE). Using a FRET-based c-di-GMP biosensor, we are able to demonstrate two roles of this incoherent feedforward loop in c-di-GMP dynamics. During growth phase transition, we show that FliA contributes to rapid regulation of c-di-GMP. Under constant growth condition, we discover remarkably rich dynamics of c-di-GMP; a subpopulation displayed large c-di-GMP fluctuations that lasted for hours, spanning multiple generations. These fluctuations are driven by pulses in flagellar gene expression, which is propagated through PdeH and RdcA. These pulsatile behaviours give rise to c-di-GMP heterogeneity in a clonal population.

Expression dynamics and localization of non-ribosomal peptide synthetases in Xenorhabdus nematophila

Although natural products (NPs) are widely studied as bioactive compounds, little is known about the expression dynamics, in vivo localization and protein abundance of the NP-producing enzymes: non-ribosomal peptide synthetases (NRPS) and polyketide synthases. Here, we used reporter gene fusions and fluorescence microscopy to study the expression dynamics and localization of NRPS. We investigated at which growth phase NRPS are expressed and in which copy numbers, as well as if they are expressed homogeneously or heterogeneously. Moreover, we studied if NRPS are spatially organized on the subcellular level.