The Role Of SPDEF In Cell Proliferation Of Gastric Cancer And The Regulatory Mechanism By Forming A Positive Regulation Loop With FoxM1

Objective:SAM-pointed domain-containing ETS transcription factor(SPDEF)is an epithelial-specific transcription factor of the E26 transformation specific(ETS)family,which binds with the target gene through the high affinity sequence of GGAT.SPDEF has been reported involved in the development of breast,prostate,and colon tumors.It is suggested that SPDEF targets the promoter activity of Forkhead Box M1(FoxM1),which has been approved by us highly expressed in gastric cancer and one of the key positive transcription factors in gastric carcinogenesis.Therefore,this study focused on the role of SPDEF in gastric carcinogenesisand the significance of the regulation mechanism on FoxM1.Methods:The gastric cancerous and matched normal gastric specimens from 22 patients were analyzed for SPDEF and FoxM1 expression using quantitative reverse transcription-polymerase chain reaction(QRT-PCR)and immunohistochemistry(IHC).The tissue microarrays were used to check the expression of SPDEF by IHC.Gene expression was assessed by QRT-PCR and Western blot depleted with small interference RNA(siRNA)or overexpressed with the special plasmid.Clonogenesis was detected by foci formation.Promoter activity was determined by dual-luciferase reporter assay.The nude mice were used as animal models to determine the ability of gastric carcinogenesis in different cell groups.Data of results were expressed as mean ± SEM and analyzed using one-way analysis of variance or Student’s t-test.Results:We investigated the expression of SPDEF in human gastric cancer species.It was determined that SPDEF was overexpressed both at the messenger RNA(mRNA)and protein levels in patients samples.The gastric cancer cells transfected with SPDEF expression plasmid or SPDEF siRNA showed the promotion or the inhibition of gastric cancer cells proliferation respectively by the clone genetics assay.Both the mRNA and protein levels of FoxM1 were regulated by SPDEF in gastric cancer cells.At the same time,FoxM1 was also overexpressed in the corresponding human gastric cancer species.The overexpression and inhibition of FoxM1 could upregulated and downregulated the mRNA and protein levels of SPDEF expression,respectively.The recovery experiments verified that the overexpression of FoxM1 could at least partially recover both the expression of SPDEF and the proliferation of the cell lines even with the siRNA inhibition of SPDEF.The result of dual luciferase activity assay showed that SPDEF bound to the promoter of FoxM1 and activated it,while FoxMl might also bind to the promoter of SPDEF to affect its expression.In the end,the results were checked in vivo.Both the inhibition of SPDEF and FoxMl reduced the growth of tumors.In this process,the two genes interacted with each other at both mRNA and protein levels.Conclusion:SPDEF is overexpressed in gastric cancer which can form a positive regulation loop with FoxM1 to promote gastric carcinogenesis.