Analysis Of Acetylome And Phosphoproteome Of Small Intestine In Domestic Cats Following Toxoplasma Gondii Infection

Toxoplasma gondii is an obligate intracellular parasite of the Phylum Apicomplexa.Approximately,one-third of the world’s population has been infected by this parasite.Generally,infection of T.gondii in people with competent immune status will not cause severe disease.However,infection of T.gondii in fetus or immunocompromised individuals will cause severe diseases and even death.T.gondii has a wide range of intermediate hosts,and almost all warmblooded animals can be the intermediate hosts of T.gnodii.However,felids are the only definitive host of this parasite.Cats play an important role in the transmission and spreading of T.gondii,and one infected cat can discharge millions of T.gondii oocysts.Intermediate hosts can be infected through ingesting food and water that have been contaminated by T.gondii oocysts.T.gondii oocysts develop in epithelial cells of feline small intestine,and protein acetylation and phosphorylation are both important post-translational modifications,thus,understanding how T.gondii reprograms the protein acetylation or phosphorylation status in feline small intestine will lay a foundation for uncovering how T.gondii interacts with its definitive host,and the mechanism underlying how T.gondii oocysts are developed.Objectives: This project aimed to study the acetylation and phosphorylation proteomic alterations of feline small intestine during T.gondii oocyst development,uncovering the impacts of T.gondii infection on protein modification in feline small intestine.Methods: In this study,the label-free quantitative acetyl-proteomic approach was used to study the acetylation alteration of proteins in feline small intestine during T.gondii oocyst development,and i TRAQ-based phosphoproteomic approach was used to study the phosphorylation alteration of proteins in feline small intestine during T.gondii oocyst development.Results: Our study showed that T.gondii infection induced significant acetylation alteration in proteins of the cat small intestine.2543 acetylated peptides and 2606 unique lysine acetylation sites were identified in 1357 acetylated proteins.The acetylation level of 334 peptides was downregulated,while 82 peptides showed increased acetylation level in the feline small intestine infected by T.gondii.Subcellular location analysis showed that most altered acetylated proteins were located in cytoplasm,nucleus,mitochondria and extracellular components.Function enrichment analysis showed that the altered acetylated proteins were enriched in glycolysis/gluconeogenesis,citrate cycle and peroxisome.Also,we identified a list of novel acetylated proteins of T.gondii,including one hypothetical protein that was expressed only in the oocysts or feline enterocytes.In phosphoproteomic analysis,a total of 4998 phosphorylated peptides and 3497 unique phosphorylated sites in 1805 phosphorylated proteins were identified.82 proteins showed down-regulated phosphorylation level,while 1289 proteins showed upregulated phosphorylation level.The functional enrichment analysis of differentially phosphorylated proteins showed that the altered phosphorylated proteins participated in actin cytoskeleton recombination,necrosis and immune response.Conclusion: This study showed that the T.gondii infection has extensive influence on the post-transcriptional modification of proteins in cat small intestine.Through this study,we have got a preliminary understanding of how T.gondii reprograms acetylation or phosphorylation modification status of definitive host proteins.Our data provided new and valuable resource for further investigating the role of protein modification in the interaction between feline small intestine and T.gondii.