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Here, we characterize the transcriptional responses of\u00a0Escherichia coli MG1655 to excess ionic copper using RNA sequencing and a genome-wide GFP-based promoter library. We applied 2\u00a0mM copper, which slows growth, and 8\u00a0mM copper, a near-lethal concentration. RNA-seq revealed extensive transcriptome remodeling, with 487 genes upregulated at 2\u00a0mM and 364 at 8\u00a0mM. Both concentrations strongly induced canonical copper-responsive systems, oxidative stress defenses, histidine biosynthesis, and multiple iron acquisition pathways\u2014including enterobactin biosynthesis and transport\u2014despite external iron failing to reduce copper toxicity. At 2\u00a0mM copper, additional pathways were activated, including heat-shock and protein-folding functions as well as lipid A, methionine and arginine biosynthesis. Copper exposure also repressed large gene sets: 486 genes at 2\u00a0mM, enriched for biofilm formation and pH elevation, and 217 genes at 8\u00a0mM, enriched for anaerobic metabolism. In contrast to the robust RNA-seq results, we investigated the Horizon Discovery E. coli genome-wide GFP based promoter library as an alternative screening tool. However, in our experiments it showed low signal-to-noise ratios, limiting its suitability for large scale gene expression screening.<\/jats:p>","DOI":"10.1093\/femsmc\/xtag050","type":"journal-article","created":{"date-parts":[[2026,9,17]],"date-time":"2026-09-17T12:42:04Z","timestamp":1789648924000},"source":"Crossref","is-referenced-by-count":0,"title":["Copper stress upregulates oxidative stress response, histidine production, and iron acquisition genes in\n                    <i>Escherichia\u00a0coli<\/i>"],"prefix":"10.1093","volume":"7","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-3340-9120","authenticated-orcid":false,"given":"Hanna","family":"Ainelo","sequence":"first","affiliation":[{"name":"Institute of Molecular and Cell Biology, University of Tartu , 51010 Tartu ,","place":["Estonia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Karola","family":"J\u00f5earu","sequence":"additional","affiliation":[{"name":"Institute of Molecular and Cell Biology, 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