Research And Application Of RNA-seq Technology In Regulation Of Host Gene Expression By Toxoplasma Gondii ROP17 And ROP18

Toxoplasma gondii is an obligate intracellular pathogen which can infect all warm-blooded vertebrates.Approximately one third of the world human population has been infected by this parasite.The apical complex of T.gondii secretes a group of rhoptry-secreted kinases(ROPs),which play significant roles in promoting intracellular infection.As crucial virulence factors of T.gondii,studies have shown that the ROP17 and ROP18 proteins of T.gondii genotype I contributes to protect T.gondii from being eliminated in interferon-activated macrophages through phosphorylating IRGs and GBPs,and then inhibiting accumulation of IRGs and GBPs and action at the parasitophorous vacuole membrane(PVM).In addition,ROP17 and ROP18 reguate host gene expression.For example,ROP18 phosphorylates p65 at Ser-468 to promote its ubiquitin-dependent degradation,which prevent the nuclear translocation of p65 and induces the NF-κB pathway termination.However,it is yet to know how ROP17 and ROP18 of T.gondii genotype I regulate host gene exression globly,and how to mediate immune evasion.Therefore,the objectives of the present study were to examine the transcriptomic changes in HEK293 T cells after transfected with PCMV-N-HA-ROP17 and PCMV-N-HA-ROP18 recombinant plasmids.Transcriptomic analysis revealed 3138 differentially expressed genes(DEGs)in PCMV-N-HA-ROP17-transfected HEK293 T cells,including 1456 upregulated,1682 down-regulated DEGs.Also,715 of the DEGs were transcription factors(TFs).Including 423 down-regulated TFs and 292 upregulated TFs,299 enrichment pathways,regardless of cancer-related signaling pathways or immune-related signaling pathways are mostly down-regulated.In the HEK293 T cells transfected with PCMVN-HA-ROP18 recombinant plasmid,1193 genes were identified as DEGs,620 DEGs were upregulated,and 573 DEGs were down-regulated.144 differentially expressed TFs were identified in this study,including 75 upregulated TFs and 69 down-regulated TFs.234 pathways were enriched,most pathways were cancer-related signaling pathways and immune system related signaling pathway that they are showing down-regulation.Taken together,this study,for the first time,explored the transcriptome changes of HEK293 T cells transfected with PCMV-HA-ROP17 plasmid and PCMV-HA-ROP18 plasmid,and reveal that ROP17 and ROP18 of T.gondii genotype I manipulates various cellular processes including immune response through reprogramming host gene expression to promote its own colonization and survival in the infected host cells.Our findings have important implications for better understanding the replication and immune evasion mechanism of T.gondii,and may also provides base-line data support for future research and development of Toxoplasma vaccines.