Integrating tissue proteomics and bioactive lipid analysis of ovine tissue samples identifies functional relations of bioactive lipids

Bioactive lipids are potent effector molecules that may help to reveal specific information about biological processes. However, analytical challenges and incomplete functional annotations remain as obstacles. We hypothesized that the functional annotation of these lipids may be improved by projecting enriched Gene Ontology (GO) terms derived from proteomics to correlating bioactive lipids. To that aim, thirteen organs and tissues from sheep ( Ovis aries ) were analyzed using mass spectrometry-based untargeted proteomics and bioactive lipid analysis. In total, 4717 proteins and 168 bioactive lipids, including fatty acids and their conjugates, oxylipins (octadecanoids, eicosanoids, and docosanoids), lyso-phosphatidylcholines (LPCs) and lyso-phosphatidylethanolamines (LPEs), endocannabinoids and related fatty amides, and bile acids and related sterol lipids were determined. Thereof, the correct identification of 114 lipids was confirmed using commercially available analytical standards, whereas the identity of the previously uncharacterized oxylipin 4-hydroxy-eicosatetraenoic acid (4-HETE) was verified through in-house chemical synthesis. Co-expression and clustering analyses are consistent with our hypothesis via independently corroborating known bioactive lipid functions and suggested novel functional relations. Amongst other observations, 18-HETE and 19-HETE were related to mitochondrial fatty acid beta-oxidation, 11,12-dihydroxy-eicosatetraenoic acid (DiHETE), 15-deoxy-prostaglandin J2 and 5(6)-epoxy-eicosatrienoic acid (EET) were related to chemical synaptic transmission, arachidonoyl ethanolamide phosphate was related to muscle contraction and numerous LPCs and LPEs were related to exosome formation. Tissue levels of polyunsaturated fatty acids (PUFAs), pro-resolving mediators and the newly characterized 4-HETE were related to the cellular protein synthesis and folding machinery. The latter finding suggested that PUFAs and their derivatives support protein synthesis fidelity in addition to efferocytosis, both essential contributors for the effective resolution of inflammation.