Mechanism Of DNMT1 / Camk2n1 Feedback Regulation And Its Mediated Methylation In Prostate Cancer

Objective: To investigate the feedback regulation of CAMK2N1/DNMT1 and the mechanism of its mediated methylation modification in prostate cancerMethods: The Cp G island distribution in the promoter and the first exon of camk2n1 gene was analyzed by Cp G island prediction software.The cells were selected to detect the DNA methylation in the promoter of camk2n1 gene by BSP method.The genomic DNA was extracted from prostate cancer and benign prostatic hyperplasia,and then the CG site of camk2n1 gene was methylated by pyrosequencing.The correlation between camk2n1 gene and DNMT1 in prostate cancer cells was analyzed by searching public databases.The DNA methylation level of camk2n1 gene promoter was detected by MSP.The knockout and overexpression plasmids targeting camk2n1 were transfected in LNCa P and DU145 cell lines of prostate cancer.The expression of DNMT1,camk2n1 and related pathway proteins were detected by RT-PCR and Western blot.Akt and ERK inhibitors were added to the above cell lines.Meanwhile,si RNA targeting camk2n1 was transfected.Results: The expression of camk2n1 gene was decreased in prostate cancer cells,and there were dense Cp G islands in its promoter region.BSP showed that compared with normal prostate epithelial cells,prostate cancer cells had hypermethylation,and thishypermethylation level was coherent to its induced down regulation of gene expression.The methylation degree of camk2n1 in prostate cancer tissue was significantly higher than that in hyperplasia tissue,and the methylation degree increased with the progress of clinical stage.Compared with BPH and PCA,the expression of camk2n1 and DNMT1 in metastatic prostate cancer also decreased.After knockout of DNMT1,the m RNA and protein expression of camk2n1 increased,while the protein expression of camk2n1 decreased after overexpression of DNMT1.After knockout of camk2n1,the expression of DNMT1 m RNA and protein increased,and the expression of p-Akt,p-mek and p-ERK also increased,while the expression of camk2n1 was opposite.Conclusion: The Cp G island in the promoter region of camk2n1 in prostate cancer cells is hypermethylation,which can induce the silence of the corresponding gene expression.Moreover,DNMT1 is the key factor affecting the methylation level of camk2n1,and camk2n1 can inhibit the expression of DNMT1 through MAPK / MEK / ERK signal pathway negative feedback.