Characterization of CDK5RAP3 in hepatocellular carcinoma

Abstract

CDK5 regulatory subunit associated protein 3 (CDK5RAP3, also called C53 and LZAP) is a novel cyclin-dependent kinase 5 (CDK5) activator interacting protein that has been shown to regulate apoptosis induced by genotoxic stress. More recently, CDK5RAP3 has also been found to regulate p53 and NF-κB signaling. Here, we have characterized the role of CDK5RAP3 in hepatocellular carcinoma (HCC) cell lines. Using real-time quantitative RT-PCR, we found that 58% (39/67) of human HCC samples had significantly higher expression (at least 2-fold) of CDK5RAP3 transcripts as compared to their corresponding nontumorous livers. The overexpression of CDK5RAP3 was statistically correlated with clinicopathological feature of more metastatic phenotype. Using stable expression clones, we demonstrated that knockdown of CDK5RAP3 in HCC cell line inhibited anchorage-independent growth and tumorigenicity of cells while overexpression of CDK5RAP3 remarkably promoted tumorigenicity of cells. Our western blotting data indicated that knockdown of CDK5RAP3 in HCC cell line SMMC-7721 resulted in an upregulation of p14ARF. Using luciferase reporter assay, we showed that ectopic expression of CDK5RAP3 suppressed p14ARF promoter activity in a dose-dependent manner. Taken together, our data provide the first evidence that CDK5RAP3 is overexpressed in HCC and overexpression of CDK5RAP3 may contribute to the development of HCC via downregulation of p14ARF. In summary, CDK5RAP3 is a potential oncoprotein in promoting hepatocarcinogenesis, the regulation behind may attribute to its regulation on p14ARF expression.