Study On The Role And Mechanism Of DEPDC1 And Its Downstream Gene CYP27B1 In Oral Squamous Cell Carcinoma

Oral squamous cell carcinoma(OSCC)is a prevalent and malignant cancer.However,the molecular mechanism of OSCC progression is not fully understood.Here,we found that the DEP domain containing 1(DEPDC1)protein was overexpressed in OSCC tissues and that the increased expression of DEPDC1 was closely associated with tumor size and poor clinical outcomes in OSCC patients.We found CYP27B1(cytochrome P450 family 27 subfamily B member),a downstream gene of DEPDC1,by m RNA sequencing of OSCC cells that overexpressed and knocked down DEPDC1.Functional investigations demonstrated that DEPDC1 stimulates OSCC cell proliferation by inhibiting CYP27B1 expression.Further,we found that upregulated expression of DEPDC1 was closely associated with smoking status in OSCC patients.In vitro experiments showed that tobacco compound NNK stimulates DEPDC1 expression by promoting the methylation of its gene body by increasing DNMT1 expression in OSCC cells.Notably,the silencing of DEPDC1 dramatically inhibited OSCC growth by inhibiting cell proliferation and inducing apoptosis in vivo.These findings suggest that smoking causes DEPDC1 overexpression in OSCC through DNMT1-regulated DNA methylation and that upregulated DEPDC1 stimulates OSCC cell proliferation by inhibiting CYP27B1 expression.Our results establish a new mechanism of OSCC progression and highlight DEPDC1 as a candidate prognostic biomarker and therapeutic target in OSCC.It provides a new experimental basis for the further study of the biological characteristics of OSCC and a new idea for the clinical treatment of OSCC.Methods1.Immunohistochemical(IHC),q RT-PCR and Western blot(WB)were used to detect the expression of DEPDC1 in the clinical samples of OSCC and the cell lines of OSCC.2.Celigo and CCK-8 were used to detect the effects of DEPDC1 on proliferation of OSCC cells,and flow cytometry detected the effect of DEPDC1 on apoptosis of OSCC cells.3.Subcutaneous and in situ tumorigenesis in nude mice.4.q RT-PCR,Western blot(WB),immunohistochemical(IHC)were used to detect the expression of CYP27B1 in the clinical samples of OSCC and the cell lines of OSCC.5.CCK-8 and Ed U were used to detect the effects of DEPDC1 and CYP27B1 on proliferation of OSCC cells.6.BSP was used to detect changes in DNA methylation after NNK-treated OSCC cells.QRT-PCR and Western blot(WB)were used to detect the expression of DNMT1 after NNK-treated OSCC cells.And western blot(WB)was used to detect the expression of DNMT1,DEPDC1,CYP27B1.Results1.DEPDC1 expression was significantly increased in OSCC tissues and closely correlated with poor clinical outcomes in OSCC patients.At the same time,the expression of DEPDC1 was also detected in OSCC cell lines.2.DEPDC1 promoted OSCC cell proliferation and inhibited apoptosis.3.DEPDC1 promoted the growth of OSCC in nude mice.4.Expression of CYP27B1 was detected in OSCC tissues and cell lines.Meanwhile,we found that CYP27B1 was negatively regulated by DEPDC1 in OSCC cells.5.CYP27B1 inhibited the proliferation of OSCC cells,and DEPDC1 promoted the proliferation of OSCC cells by inhibiting the expression of CYP27B1.6.NNK stimulated DEPDC1 expression through DNMT1 in OSCC cells.Conclusions1.DEPDC1 plays a carcinogenic role in OSCC.2.DEPDC1 plays a carcinogenic role by inhibiting the expression of CYP27B1 in OSCC.3.NNK induced DEPDC1 up-regulation by promoting DNMT1 expression in OSCC.