The Mechanism On Transcription Factor C-Jun Alters Invasion And Migration Abilities Of Liver Cancer Cells By Regulation Of β3GnT8

Objective: To investigate the transcriptional regulation of glycosyltransferase β3GnT8 by transcription factor c-Jun and the research on transcription factor c-Jun and β3GnT8 in invasion and metastasis capability of liver cancer cells.(1) Real time PCR and Western blot were conducted to detect c-jun and β3GnT8 expression in SK-Hep-1, SMMC-7721 and HepG2 cell lines;(2) The interaction of transcription factor c-Jun and promoter of β3GnT8 was detected by using ChIP assay;(3) Exogenous c-jun sense plasmid vector was transfected into SK-Hep-1 cells and c-jun shRNA interference vector was transfected into HepG2 cells by lipofectamine. Then, transfection efficiency was detected by fluorescence microscope;(4) The mRNA and protein expression of c-Jun, β3GnT8, and CD147 were detected by Real time PCR and Western blot;(5) Lectin blot was used to assess the expression of polylactosamine;(6) Transwell migration and invasion analysis were conducted to examine the change of potential of migration and invasion in the two groups;(7) Cell scratch repair assessment was used to detect the migration ability of the two groups;(8) β3GnT8 stable up-regulated cell lines were established;(9) c-jun shRNA interference vector was transfected into SK-Hep-1 and SMMC7721 β3GnT8 up-regulated cell lines, then expression of c-Jun, β3GnT8, CD147, polylactosamine and potential of migration and invasion was analyzed.Results:(1) Among the cell lines of HepG2, SMMC7721 and SK-Hep-1, HepG2 expresses the highest level of c-Jun and β3GnT8. SK-Hep-1 expresses the lowest level of c-Jun, and β3GnT8 expression is low in SMMC7721;(2) ChIP assay shows that activated c-Jun protein is able to combine to promoter fragment of β3GnT8;(3) The expression of β3GnT8 and HG-CD147 were upregulated with the overexpression of c-jun, however, when c-jun was down regulated, β3GnT8 and HG-CD147 expression were significantly downregulated;(4) Lectin blot analysis shows that polylactosamine expression of the two cell lines was changed following the c-jun overexpression or downregulation;(5) The potential of migration and invasion of SK-Hep-1 were enhanced when c-jun was up regulated; In HepG2 cells, migration and invasion abilities were declined following the interference expression of c-jun;(6) In SK-Hep-1 and SMMC7721 β3GnT8 up-regulated cell lines, compared to the control, the expression of β3GnT8, HG-CD147, polylactosamine and potential of migration and invasion were totally increased. When c-jun was down regulated in SK-Hep-1 and SMMC7721 β3GnT8 up-regulated cell lines, the expression of β3GnT8, CD147, polylactosamine and potential of migration and invasion were also down regulated in these cell lines.Conclusion:(1) β3GnT8 plays an essential role in the invasion and migration of liver cancer by synthesis the formation of polylactosamine and changing level of glycosylation of CD147. Overexpression of β3GnT8 can enhance the potential of invasion and migration of SK-Hep-1 and SMMC7721 cell lines;(2) Transcription factor c-Jun can alter cell migration and invasion of liver cancer. And the regulation of β3GnT8 by c-Jun may be one of the mechanisms that c-Jun regulates the migration and invasion of liver cancer.