Conclusions
This study presents a novel experimental design combined with an innovative research tool for systems-level analyses of molecular phenotypes (DIA-LCMS2). Specific organismal phenotypes (blood osmolality, body condition, feeding rate) were found to correlate well with extreme salinity stress in O. mossambicus . The rate-independent salinity maximum and critical threshold salinity were determined using a comprehensive set of acclimations and a generalized protocol that is reproducible for other species. The DIA-LCMS2 approach was shown to reveal comprehensive molecular phenotypes in ecologically relevant samples. The data generated in this study were used to produce protein networks which elucidate the mechanisms of ionic and mitochondrial protein regulation in response to hypersalinity in general and with respect to the critical threshold. These networks reveal novel information regarding the modulation of energy metabolism, cell adhesion, and cell structure, which includes functional annotation of previously uncharacterized proteins.