Thermal Shifts Dominate Single‐Gene Responses to Transcription‐Targeting Antibiotics

Abstract

ABSTRACT Combined exposure to thermal and antibiotic stresses is ubiquitous. How cells adapt to these concurrent cues at the single‐gene level remains unresolved. Here, we show that in Escherichia coli, both cold and heat shock responses dominate antibiotic stress responses across multiple levels of physiology. Using RNA‐seq, we demonstrate that combined exposure produces transcriptomes that are largely indistinguishable from those induced by thermal shifts alone. Flow‐cytometry measurements across intermediate temperatures and increasing ofloxacin concentrations further show that temperature‐like responses are more prevalent outside the 30°C–39°C range. This dominance arises from temperature‐driven changes in ATP levels and nucleoid organization, which in turn modulate DNA gyrase and RNA polymerase engagement with DNA. Combined exposure also induces SoxS‐ and MarA‐mediated efflux systems more strongly than individual stresses. Machine learning analysis identifies CRP and σ38 as key gene expression regulators under combined stress. Molecular dynamics simulations show that drug‐target conformational plasticity is altered under combined conditions. Cross‐species simulations across six bacterial species with distinct pathogenic profiles reveal that thermal modulation of antibiotic response is partially conserved across evolutionarily distant species. Together, the results identify temperature as a critical determinant of antibiotic response, suggesting that thermal shifts may facilitate bacterial adaptation to antibiotics.