Skin proteomic screening and functional analysis of differential proteins associated with coat color in sheep (Ovis aries)

Abstract

Objective: Coat color is an important characteristic and economic trait in domestic sheep. In this study, we explored the potential mechanisms and the signaling pathways involved in coat color regulation for sheep.Methods: Isobaric tags for relative and absolute quantification (iTRAQ) technology was used to catalog global protein expression profiles in skin of sheep with black versus white coat color. Immunofluorescence was used to observe the expression localization of differential protein. Western blot and quantitative real time polymerase chain reaction (qRT-PCR) were used to evaluate their role in the coat color formation of sheep.Results: A total of 136 differential proteins were obtained in different coat colors, including 101 up-regulated and 35 down-regulated. Pigmentation function entries were enriched through gene ontology annotation. Tyrosine metabolism and platelet activation signaling pathway were extracted by Kyoto encyclopedia of genes and genomes analysis. Apolipoprotein A-1 (APOA1) and fibrinogen alpha chain (FGA) were found to be critical differential proteins by the interaction of differential proteins in the direct-interaction network diagram. Strikingly, twenty candidate differential proteins were screened, from which beta-actin (ACTB) protein showed higher expression in white sheep skin, while albumin (ALB), APOA1, MAOA (amine oxidase) and FGA proteins showed higher expression in black sheep skin, which was validated by immunofluorescence, western blot, and qRT-PCR.Conclusion: This study identified several novel proteins that may be involved in the coat color formation of sheep. The white and black sheep skin proteome profiles obtained provide a valuable resource for future research to understand the network of protein expression controlling skin physiology and melanogenesis in sheep.