Updated project metadata. Updated publication reference for PubMed record(s): 30095222. The transcriptome of the ecotoxicological model Enchytraeus crypticus is well studied but the downstream changes at the protein level remained a gap. Changes in the protein regulation following exposure to CuO nanomaterial (NM) and Cu salt (CuCl2) were investigated. High performance liquid chromatography with tandem mass spectrometry using tandem mass tags was used. CuO NM elicited higher number of differentially expressed proteins compared to CuCl2 with little to no overlap of proteins. CuO NM caused more stress response mechanisms, with good agreement between differentially expressed proteins, genes and metabolites. CuCl2 caused higher impact in shorter time periods, but organisms have conserved mechanisms (constitutive genes) that allow Cu handling and detoxification. CuO NM caused higher impact after a longer exposure period, inducing regulation of facultative genes with a whole differentiated paradigm and cascade. This could be due to different issues: 1) the cell uptake route is different for Cu NM and Cu ions 2) internalized Cu NM can result in a “Trojan- horse” effect 3) the cascade of events occurs in a different time order 4) the organism uptake is different between life stages, i.e., cocoons thickened surface protects the entry of NM and juveniles have facilitated entry via tegument. Protein responses are of key importance when trying to understand the link between exposure and the related adverse biological effects. We here used advanced proteomic profiling techniques to describe how the protein responses differ when an organism is exposed to Copper (Cu) nanomaterial compared to when it is exposure to Cu ions. We observed that when organisms are exposed to the nano-form they have longer-term changes in protein patterns, patterns that are different from the responses following exposure to the ionic form. On the level of individual proteins we identified nano-specific mechanisms.