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Abstract Salinity tolerance in fish involves a suite of physiological changes, but a cohesive theory leading to a mechanistic understanding at the organismal level is lacking. To examine the potential of adapting energy homeostasis theory in the context of salinity stress in teleost fish,Oreochromis mossambicuswere acclimated to hypersalinity at multiple rates and durations to determine salinity ranges of tolerance and resistance. Over 3000 proteins were quantified simultaneously to analyze molecular phenotypes associated with hypersalinity. A species‐ and tissue‐specific data‐independent acquisition (DIA) assay library of MSMS spectra was created. Protein networks representing complex molecular phenotypes associated with salinity acclimation were generated.O. mossambicushas a wide “zone of resistance” from 75 g/kg salinity to 120 g/kg. Crossing into the zone of resistance resulted in marked phenotypic changes including blood osmolality over 400 mOsm/kg, reduced body condition, and cessation of feeding. Protein networks impacted by hypersalinity consist of electron transport chain (ETC) proteins and specific osmoregulatory proteins. Cytoskeletal, cell adhesion, and extracellular matrix proteins are enriched in networks that are sensitive to the critical salinity threshold. These network analyses identify specific proteome changes that are associated with distinct zones described by energy homeostasis theory and distinguish them from general hypersalinity‐induced proteome changes. 

Acclimations of Oreochromis mossambicus to elevated salinity were conducted with multiple rates of salinity increase and duration of exposure to determine the rate-independent maximum salinity limit and the incipient lethal salinity. A data-independent acquisition (DIA) assay library was created for quantitative analysis of over 3000 gill proteins simultaneously in treatments representative of important key zones in the salinity level/duration landscape. From these DIA data, protein networks that represent complex molecular phenotypes associated with salinity acclimation were generated. Acclimations of Oreochromis mossambicus to elevated salinity were conducted with multiple rates of salinity increase and duration of exposure to determine the rate-independent maximum salinity limit and the incipient lethal salinity. A data-independent acquisition (DIA) assay library was created for quantitative analysis of over 3000 gill proteins simultaneously in treatments representative of important key zones in the salinity level/duration landscape. From these DIA data, protein networks that represent complex molecular phenotypes associated with salinity acclimation were generated. Protein fold change (FC) and organismal level performance indicators of salinity tolerance were then correlated. Gill protein networks impacted at extreme salinity levels both above and below the incipient lethal limit include increased energy metabolism, especially upregulation of electron transport chain complex proteins, and significant downregulation of a previously uncharacterized protein which bears strong amino acid sequence similarity to fucolectin. Proteins networks strongly impacted by crossing into the zone of resistance include cell adhesion and extracellular matrix regulation.