Understanding how environmental sugars influence intracellular pH (pHi) is crucial, as pHi homeostasis can be impacted through biofilm formation in Escherichia coli. Biofilms—structured communities of bacteria attached to surfaces—protect the cells from environmental stressors, including desiccation and exposure to antibiotics. Altered pHi regulation within biofilms may be able affect bacterial metabolism, stress response pathways, and even the efficacy of antibiotics. In this project, we examined whether the presence of L-arabinose, a common pentose, affects the pHi of E. coli and whether the cells maintain pH homeostasis under these conditions. Our lab's previous transcriptomic data indicate significant differences in the expression of acid stress response genes when L-arabinose is present in the medium. To explore the potential impact of L-arabinose on intracellular pH regulation, we utilized E. coli strain PHL628 containing the plasmid pS2513.PHP which expresses a pH-sensitive GFP, modified to also contain a chloramphenicol resistance cassette for selection. By monitoring the GFP fluorescence, we tracked changes in intracellular pH in response to arabinose.

These data complement our transcriptomic data for PHL628 biofilm and planktonic cells in the presence of arabinose, which showed the activation of pH-responsive stress pathways and efflux pumps, among other changes to metabolic pathways. This protocol enables us to compare intracellular acidification in arabinose-treated and untreated cells, providing insights into the role of araE, an arabinose symporter, in pH regulation and the metabolic response to L-arabinose. Our findings shed light on how E. coli adjusts its internal pH under varying environmental conditions, with implications for biofilm formation and antibiotic resistance.

Jenna Frizzell

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