Integrated multi-omics analysis reveals divergent molecular responses in Palmer amaranth ( Amaranthus palmeri ) biotypes susceptible and resistant to glyphosate

Environmental stress triggers coordinated changes across genetic, transcriptomic, proteomic, and metabolomic levels in plants, yet the extent of synchronization across these omic layers remains underexplored. We captured transcriptomic, proteomic and metabolomic perturbation of glyphosate-resistant (GR) and glyphosate-susceptible (GS) Palmer amaranth ( Amaranthus palmeri ) biotypes 24 hours after herbicide treatment, quantifying 30,371 transcripts, 5,606 proteins, and 220 metabolites. Glyphosate perturbed threefold more transcripts and proteins in GS than in GR and caused the accumulation of shikimate intermediates in both biotypes. In GS, glyphosate severely disrupted primary metabolism, including photosynthesis and carbon fixation, leading to a collapse of energy production and impairment of phenylpropanoid and terpenoid biosynthesis, compromising defense and detoxification. In contrast, GR maintained cellular homeostasis, with minimal perturbation in carbon metabolism and upregulation of detoxifying pathways, indicating metabolic rerouting. Integrated multi-omics analysis captured stress responses hidden from single-omic analysis, including elevated glutathione metabolism, perturbation of the phenylpropanoid pathway and elevated raffinose family oligosaccharide metabolism in GR, and perturbation of taurine-hypotaurine metabolism in GS. Transcript and protein changes were broadly correlated, but GS exhibited signs of translational inhibition under glyphosate stress, indicating reduced protein synthesis. These findings reveal pervasive perturbation of glyphosate beyond the shikimate pathway within 24 hours after exposure, and underscore the importance of multi-omics integration to elucidate complex stress responses in plants.