Identification and characterization of the human Cdc2l2 gene promoter

The CDK11 (cyclin-dependent kinase 11, formerly known as PITSLRE) protein kinases are part of the large family of p34^cdc2-related kinases and have been shown to play a role in cell cycle progression, RNA processing and apoptosis. They are encoded by two genes - cell division control like 1 (Cdc2L1) and cell division control like 2 (Cdc2L2). To date, little is known about the transcription factors controlling their expression. To understand the mechanisms underlying the regulation of CDK11 gene expression, we cloned and identified the Cdc2L2 promoter and determined its transcriptional regulatory elements. By deletion analysis, a region between nucleotides -145 and +10 was identified to be critical for basal level transcription of the Cdc2L2 gene. Sequencing analysis revealed that the proximal promoter of the Cdc2L2 gene is GC rich and does not contain TATA and CAAT boxes. However, multiple consensus and near consensus transcription factor binding sites were found to be present in this region, such as two Ets-1, one cAMP-responsive element (CRE) and one TCF11/LCR-F1/Nrf1 binding sites. Site-directed mutagenesis and transfection studies revealed that all these binding sites were necessary to achieve sustained transcriptional activity. Electrophoretic mobility shift assay confirmed that transcription factors Ets-1 and CREB bind to the Cdc2L2 promoter elements, indicating their potential role in the transcriptional regulation of Cdc2L2 gene. More importantly, Ets-1, CREB and phosphorylated CREB were found binding to the endogenous Cdc2L2 promoter using chromatin immunoprecipitation (CHIP) assay. Our results provide the foundation for further studies into the regulation of Cdc2L2 gene expression in normal homeostasis and cancer.