Phospho-proteomic analysis of the effect of merecidin inhibiting the human lung adenocarcinoma A549 cells

Abstract Background: Merecidin induced A549 cells apoptosis. The present study aimed to assess the differentially phosphorylated proteins in lung cancer A549 cells treated with antibacterial peptide merecidin. Methods: TMT/iTRAQ labeling, LC-MS/MS analysis, HPLC grading, IMAC modification and enrichment, and liquid chromatography-mass spectrometry of peptide. The data of proteins were filtered according to the localization probability >0.75 standard. The phosphorylated proteomics data were analyzed using GO, KEGG, and STRING databases. Western blots examined the changes in phosphorylated protein expression. Results: Protein function enrichment showed significant changes in the phosphorylation level with respect to protein binding, metabolic activity, molecular function regulation, cell process, and biological function regulation. Integrated pathway bioinformatics results showed that differential proteins are associated with several pathways, including mTOR and AMPK etc. Screening of the COG database revealed differentiated phosphorylated proteins in cell signal transduction, RNA transcription, translation processing and modification, ribosome synthesis of proteins, cytoskeleton protein formation, intracellular material transport, secretion, and vesicle transport. Protein interaction level analysis identified an interaction network with HDAC1, RPL23A, SRSF3H, and NCBP1.etc. The phosphorylated proteomics data and showed that after merecidin treatment, ATG2B and ULK1s were significantly upregulated, while MAPK1 and AKT were significantly downregulated. In addition, Western blot also showed the upregulated level of ATG13,ULK1;MAPK1 downregulated. Conclusion: The results verified the feasibility of phosphorylated proteomics analysis, confirmed the signaling pathways, and suggested that merecidin may induce autophagy of lung adenocarcinoma A549 cells.