To Explore The Effect Of HPV E6 And E7 Protein On Gene Expression And Autophagy Level In Host Cells

Human papillomaviruses(HPVs)are small double-stranded DNA viruses that include more than 200 genotypes.Based on differences in their pathogenicity,the HPV family can be further classified into low-risk HPV(LR-HPVs)types and high-risk HPV(HR-HPVs)types.HR-HPVs are associated with some tumors,such as cervical cancer,anal cancer.And LR-HPVs are often etiologically associated with benign infectious diseases,such as genital warts.Due to the infectious and proliferative nature of HPV,HPV related diseases have become a huge impact on the patient's physical health and a serious threat to public economic development.Therefore,research on the pathogenesis and treatments of HPV have always been an important aspect in contemporary.Numerous studies have revealed that HPV E6 and E7 play an important role in the pathogenic process during HPV infection,but their pathogenic mechanisms need to be further explored.Although E6 and E7 are both early proteins that are co-expressed during the course of HPV infection,the dissection of their individual contribution is important for determining their specific functions.This study investigated the gene expression profiles and gene expression differences of HPV 6b E7/HPV 11E6/HPV 16E7 stable cell line and vector controls by Gene microarray.The GO and KEGG analysis were used to classify genes that were expressed differentially between studied and control group,in order to discover the biological processes and pathways affected by HPV E6 / E7 protein.And this research used q PCR and Western blot to verify the differential expression genes between groups and explore the mechanisms.The expression of the S100A8/A9 genes was increased significantly in HPV 6b E7 cells,a finding from Gene microarray.Furthermore,the m RNA and protein levels of S100A8 / A9 was proved to be significantly increased in the HPV 6b E7-Ha Ca T cells which was confirmed by WB and q PCR.Until now,there was no study focusing on the relationship between S100A8 / A9 and low-risk HPV E7.So this finding may provide new direction to explore the function of LR-HPV E7;From the Gene microarray analysis,the HPV 11E6-expressing cells exhibited higher ATG10 protein levels than the control cells,indicating that HPV 11E6 might affect autophagy level in host cells.Western blot and immunofluorescence were used to detect the expression of autophagy-related genes.The results showed that HPV 11E6 can promote the expression of LC3B-II and Beclin-1.The m Cherry-GFP-LC3 plasmid was transfectcted into the cells to further confirm the above results.All the evidence indicated that HPV 11E6 can induce autophagy in host cells.In order to find out how HPV 11E6 affects autophagy,the m TOR pathway and its four upstream pathways were detected by Western blot.The results determined that HPV 11E6 inhibits Erk / m TOR and AKT / m TOR pathways to increase intracellular autophagy levels.The results expand our understanding of the relationship between low-risk HPV E6 protein and host cells;From the HPV 16E7 microarray analysis,the expression of ITGBL1 was elevated at the m RNA and protein levels.The results of immunohistochemistry showed that the expression of ITGBL1 in HPV 16(+)cervical cancer was also increased significantly.In vitro experiments indicated that HPV 16E7 affected the proliferation and migration by increasing the expression of ITGBL1 protein.Above all,this research explores the function and pathogenic mechanism of HPV E6 / E7 by using Gene microarray analysis,which deepening the understanding of the association between HPV and host cells.