Skin mucus of gilthead sea bream (sparus aurata l.). protein mapping and regulation in chronically stressed fish

The skin mucus of gilthead sea bream was mapped by one-dimensional gel electrophoresis followed by liquid chromatography coupled to high resolution mass spectrometry using a quadrupole time-of-flight mass analyzer. More than 2,000 proteins were identified with a protein score filter of 30. The identified proteins were represented in 418 canonical pathways of the Ingenuity Pathway software. After filtering by canonical pathway overlapping, the retained proteins were clustered in three groups. The mitochondrial cluster contained 59 proteins related to oxidative phosphorylation and mitochondrial dysfunction. The second cluster contained 79 proteins related to antigen presentation and protein ubiquitination pathways. The third cluster contained 257 proteins where proteins related to protein synthesis, cellular assembly, and epithelial integrity were over-represented. The latter group also included acute phase response signaling. In parallel, two-dimensional gel electrophoresis methodology identified six proteins spots of different protein abundance when comparing unstressed fish with chronically stressed fish in an experimental model that mimicked daily farming activities. The major changes were associated with a higher abundance of cytokeratin 8 in the skin mucus proteome of stressed fish, which was confirmed by immunoblotting. Thus, the increased abundance of markers of skin epithelial turnover results in a promising indicator of chronic stress in fish.
This study was funded by the European Union (AQUAEXCEL, FP7/2007/2013; grant agreement No. 262336, Aquaculture infrastructures for excellence in European fish research) project. The views expressed in this work are the sole responsibility of the authors and do not necessarily reflect the views of the European Commission. Additional funding was obtained from the Spanish Ministerio de Economía y Competitividad (MI2-Fish, AGL2013-48560) and from Generalitat Valenciana (PROMETEO FASE II-2014/085). Proteomics study was done at Proteomics laboratory of University of Valencia, Spain (SCSIE). This laboratory is a member of Proteored, PRB2-ISCIII and is supported by grant PT13/0001, of the PE I+D+i 2013–2016, funded by Instituto de Salud Carlos III and Fondo Europeo de Desarrollo Regional (FEDER).