MiR-218-5p/CDCP1 Axis Medicates KSHV VIRF1-nduced Endothelial Cells Migration And Invasion

Background: As an oncogenic virus,Kaposi's sarcoma-associated herpesvirus(KSHV)encodes many oncogenes,wherein open reading frame(ORF)K9 encoding viral interferon regulatory factor 1(v IRF1).v IRF1 can prevent interferon regulatory factor 3(IRF3)binding to CBP/P300 and induce the expression of IFN-?/? and IFN stimulated gene.In addition,v IRF1 has been reported to promote the transformation of NIH3T3 cells.Our previous studies have found that overexpression of v IRF1 in human umbilical vein endothelial cells(HUVEC)could induce cell proliferation,migration,invasion and microtubule formation.With isobaric tags for relative and absolute quantitation(i TRAQ)technology,the altered expressions of many proteins have been indicated,including CUB domain-containing protein 1(CDCP1).Objective: To explore the role of CDCP1 and micro RNAs(mi RNAs)in KSHV v IRF1-induced endothelial cells migration and invasion.Methods: The alterd expression of CDCP1 which was involved in the regulation of cells migration and invasion was found by i TRAQ,and the result was confirmed by real-time quantitative polymerase chain reaction(RT-q PCR),immunohistochemistry(IHC)and Western blot.Next,the recombinant lentivirus of short hairpin RNA of CDCP1 was constructed and used to infect v IRF1-overexpressing HUVEC.Then Transwell migration and invasion assays were performed in CDCP1-knockdown HUVEC.Meanwhile,microarray analysis was used to detect the altered mi RNAs in v IRF1-overexpressing HUVEC,and luciferase reporter assay and Western blot were further utilized to explore the mi RNAs targeting the 3' untranslated region(UTR)of CDCP1.more experiments were used to confirm the relationship between the candidate mi RNA and CDCP1,gain-of-function assay was applied to determine the role of the candidate mi RNA in KSHV v IRF1-induced endothelial cells migration and invasion.Results: CDCP1 was highly expressed in v IRF1-overexpressing HUVEC by realtime PCR and Western blot.More importantly CDCP1 in Kaposi's sarcoma(Kaposi's sarcoma,KS)skin tissue was significantly higher than that in normal skin tissue.The results from Transwell migration and invasion assays in CDCP1-knockdown HUVEC indicated that CDCP1 knockdown significantly reduced KSHV v IRF1-induced endothelial cells migration and invasion.The mi R-218-5p was found to target CDCP1 3'UTR by microarray analysis,luciferase reporter assay and Western blot.Further functional experiments confirmed that mi R-218-5p was involved in KSHV v IRF1-induced endothelial cells migration and invasion by targeting CDCP1.Conclusions: KSHV v IRF1 promotes endothelial cell migration and invasion by upregulating CDCP1,which is targeted by mi R-218-5p.