ABSTRACT: The present work was devoted to a multi-level characterization of E. coli exposed to Ag+-mediated stress using for the first time an approach of integrative biology, based on the combination of physiological, biochemical and transcriptomic data sets. Bacterial growth and survival after Ag+ exposure were first quantified and related to the accumulation of intracellular silver, as detected by Nano Secondary Ion Mass Spectroscopy (NanoSIMS) at high lateral resolution. The whole transcriptomic response of E. coli cells under ionic silver-mediated stress was then characterized. Clear correlations were established between (i) cell physiology, (ii) variations in the biochemical characteristics of cell fatty acids and proteins, and (iii) regulation of gene expression. This challenging approach allowed determining key genetic markers of the E. coli response to ionic silver. In particular, we identified Ag+-mediated regulations of gene expression in correlation with growth (e.g. genes of transporters, transcriptional regulators, ribosomal proteins), necessary for ionic silver transport and detoxification (e.g. copA, cueO, mgtA, nhaR) and to cope with various stress (dnaK, pspA, metA,R, oxidoreductase genes). Regulation of gene expression after Ag+ exposure was also correlated to macromolecular modifications, such as acyl chain length (e.g. fadL, lpxA, arnA), protein secondary structure (e.g. dnaJ, htpX, degP) and cell morphology (e.g. ycfS, ycbB).