Abstract: 【Objective】 This study aimed to investigate the immune response mechanism of S-adenosyl methionine domain-containing protein 2 （RSAD2） under Orf virus （ORFV） infection， providing theoretical basis for the control and treatment of Orf. 【Method】 Transcriptome sequencing was performed on goat skin fibroblast （GSF） cells （control group）， stably expressing non-targeted shRNA GSF cells （negative control group）， and RSAD2-knockdown GSF cells （RSAD2i GSF cells， stably expressing RSAD2 targeting interference shRNA ） at 12 h post ORFV infection. The sequencing data of cells from each group at 0 and 6 h post infection from previous studies were also used for comparison. 【Result】 The screening results of differentially expressed genes （DEGs） showed that RSAD2i GSF cells had 1923 DEGs at 12 h post infection compared with 0 h post infection. Gene set enrichment analysis （GSEA） indicated that RSAD2i GSF cells exhibited more significant down-regulated immune-related signaling pathways at 6 h post infection than at 12 h post infection （P<0.05，the same below）. Database for annotation， visualization， and integrated discovery （DAVID） and KEGG orthology based annotation system （KOBAS） analyses showed that RSAD2i-ORFV gene set was enriched to immune-related signaling pathways， such as IL-17 signaling pathway， cytokine-cytokine receptor interaction， and TNF signaling pathway. The 424 DEGs of RSAD2i-ORFV gene set showed 8 expression patterns at 0， 6， and 12 h post infection （R0→R6→R12） in RSAD2i GSF cells. Among them， the expression patterns 1， 2， and 5 were significant and were different types of down-regulated expression. Protein-protein interaction （PPI） network analysis indicated that IL-6， IL-1α， and IL-1β had more associations and stronger interactions with other proteins， with IL-6 being predicted as the most key protein in the PPI network. Real-time fluorescence quantitative PCR detection showed that the changing trend of relative expression of TNFSF15， IL-1B， MAP3K7， TGFB2， MAPK6， and IL-11， except IL-13RA2 were consistent with the sequencing results. 【Conclusion】 At the early stage of ORFV infection， RSAD2 may positively regulate the IL-17 signaling pathway to promote the production of downstream pro-inflammatory cytokine IL-6， thereby mediating the defense response of the host to ORFV； ORFV may achieve immune escape with the assistance of encoded proteins to directly influence the IL-17 signaling pathway. The RSAD2—IL-17—IL-6 axis may be the potential mechanism for host resistance to ORFV infection.