RUNX1 Inhibits Colorectal Tumor Growth And Metastasis Through Regulating CDK8

Colorectal cancer(CRC)is a common gastrointestinal malignant cancer worldwide,whose morbidity and mortality are still increasing.Metastasis is the leading cause of death in metastatic CRC patients,with a five-year survival rate of 13%.Upon dimerization with CBFβ,RUNX(Runt-related transcription factor),a class of highly conserved transcription factor,achieves high-affinity of DNA binding and regulates the expression of its target genes.Studies have shown that RUNX(RUNX1,RUNX2,RUNX3)plays an important role in cell proliferation and differentiation.However,the biological function and mechanism of RUNX1 in CRC still remain unclear.First,immunohistochemistry and western blot were used to detect the expression of RUNX1 in CRC tissue samples and normal adjacent tissue samples.The results showed that RUNX1 was significantly decreased in CRC tissues,suggesting that RUNX1 might play a role as a tumor suppressor gene in CRC.Next,we used ChIP-seq to analyze the transcriptional regulatory targets of RUNX1.RUNX1 antibody was used to perform chromatin immunoprecipitation assay in HCT116 cells.23 genes with an enrichment value over 25 were acquired.Among them,5 genes were significantly dysregulated in CRC cells stably over-expressing RUNX1.Furthermore,only CDK8 was significantly increased in CRC patients tissues,suggesting CDK8 may be a transcriptional regulatory target of RUNX1.And this regulating manner was next analyzed in CRC cells.ChIP-qPCR results showed that RUNX1 mainly binds to the promoter region of 499-379bp at the upstream of CDK8’s transcription initiation site.In addition,RUNX1 over-expression can down-regulate the mRNA and protein levels of CDK8,suggesting it can be transcriptionally regulated by RUNX1.Because CDK8 is related to the proliferation,migration and cell cycle of CRC cells,we detected the expression of the downstream target genes of CDK8 using qRT-PCR.The results showed that these genes were significantly dysregulated in CRC cells stably over-expressing RUNX1.By means of CCK8,cell scratch,transwell,cell cycle analysis in vitro,it was found that RUNX1 can inhibit the proliferation and migration of CRC cells and block the cell cycle transition to S phase.Furthermore,RUNX1 could inhibit CRC tumor growth and metastasis in xenograft and liver metastasis mouse models.Together,our results may help to further understand the pathogenesis of CRC,and RUNX1/CDK8 may provide novel approaches and targets for CRC prevention and treatment.