Characterization of the Immunomodulatory Mechanism of a Pleurotus eryngii Protein by Isobaric Tags for Relative and Absolute Quantitation Proteomics

PEP 1b is a novel immunoregulatory protein isolated from Pleurotus eryngii, a popular edible mushroom. In this study, isobaric tags for relative and absolute quantitation (iTRAQ) approach and bioinformatics analysis were used to characterize the PEP-1b-induced proteome alterations in Raw 264.7 macrophage cells, to comprehensively excavate the molecular mechanisms involved in the immunoregulatory effects of PEP 1b. In comparison to the control group, PEP 1b treatment significantly changed the expression of 292 proteins, including 191 upregulated and 101 downregulated proteins. Bioinformatics analysis showed that PEP-1b-regulated proteins were involved in 437 biological process domains, 131 cellular component domains, and 90 molecular function domains. Moreover, PEP 1b played the role of immunomodulator by mainly modulating the Rap1 signaling pathway, Wnt signaling pathway, Ras signaling pathway, and PI3K-Akt signaling pathway. Interestingly, PEP 1b regulated the proteins involved in the immune system, signal transduction, and transport processes, which were related to the immunoregulatory effects of PEP 1b. The western blotting analysis confirmed that the immune-boosting activities of PEP 1b were associated with modulating the expression of Sqstm1, Cox2, Rap1b, and Pyk2. The current research provided a comprehensive understanding of the immunoregulatory effects and molecular mechanisms involved in the PEP 1b supplementation.