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Proteomics plays a significant role in discerning the effects of chemical exposures in animal taxa. Multi-omics applications have become more pervasive in toxicology, however questions remain about whether proteomics is being utilized by the community to its full potential - are we placing too much stock in transcriptomics and other omics approaches for developing adverse outcome pathways? Proteins are more relevant than transcripts because they are direct mediators of the resulting phenotype. There is also rarely perfect stoichiometry between transcript and protein abundance and transcript abundance may not accurately predict physiologic response. Proteins direct all levels of phenotype: structural proteins dictate physical form, enzymes catalyze biochemical reactions, and proteins act as signaling proteins, antibodies, transporters, ion pumps, and transcription factors to control gene expression. Molecular initiating events (MIEs) of AOPs predominantly occur at the level of the protein (e.g. ligand-receptor binding) and proteomics can elucidate novel MIEs and mapping KEs in AOPs. This critical review highlights the need for proteomics in multi-omics studies in environmental toxicology and outlines steps required for inclusion and wider acceptance in chemical risk assessment. We also present case studies of multi-omics approaches that utilize proteomics and discuss some of the challenges and opportunities for proteomics in comparative ecotoxicology. Our intention is not to minimize the importance of other omics technologies, as each has strengths and limitations, but rather to encourage researchers to consider proteomics-based methods in multi-omics studies and AOP development.

Advances in proteomics have greatly improved chemical toxicity assessment and predictions of adverse outcomes in organisms. Ecotoxicoproteomics has been employed to elucidate biological pathways affected by chemicals and provide data that can be incorporated into adverse outcome pathways (AOP) to better define the ecological risk of emerging pollutants. Microplastics (MPs) and plastic additives have raised global concern due to their widespread use in aquatic environments, bioaccumulation in tissues, and toxic effects in aquatic organisms. Despite showing sublethal toxicity in many cases, mechanisms underlying these emerging pollutants are underexplored. In this review, adverse effects and recent ecotoxicoproteomic studies of MPs and typical additives (i.e. plasticizers, flame retardants, antioxidants, and UV stabilizers) in aquatic organisms are summarized. Proteomics data show that MPs adversely affect ingestion and reproduction via disrupting pathways related to energy metabolism, stress-related defense, and cytoskeletal dynamics. Biological processes including lipid metabolism, energy homeostasis, skeletal development, neurotransmitter signaling, and immune response are modulated by additives and induce developmental malformations in fish embryos/larvae. Furthermore, plastic additives also exert reproductive toxicity, hepatotoxicity, and neurotoxicity in invertebrates (e.g. mussel, abalone, and oyster) and fish by disrupting detoxification/oxidative stress, hormonal modulation, signal transduction, and apoptosis. Additional studies are needed to complement the omic knowledge of chemical additives that are not well documented (e.g. UV stabilizers) for improving understanding into toxic mechanisms and for characterizing ecological risk linked to plastic contaminants.