Expression of PDZ-binding kinase (PBK) is regulated by cell cycle-specific transcription factors E2F and CREB/ATF.
Earlier we reported that a novel mitotic protein kinase, PDZ-binding kinase (PBK), is expressed in primary hematopoietic neoplasms. Recent reports have suggested a role for PBK in mitotic progression. In the present study, we demonstrate that PBK is down regulated during doxorubicin induced growth arrest of HL60 promyelocytic leukemia cells at least partly due to cell cycle-specific transcriptional regulation. Furthermore, we show that transcriptional control is mostly due to binding of transcription factors E2F and CREB/ATF to two distinct binding sites within the PBK promoter. This was demonstrated by: (i) electrophoretic mobility shift assays showing transcription factor binding within the PBK promoter at the putative E2F (-146bp) and CREB/ATF (-312bp) binding sites; (ii) Western immunoblot analysis of knockdown extracts from siRNA inhibition of transcription factor expression showing that PBK protein expression is dependent upon the presence of these transcription factors; (iii) codistribution of CREB factor and PBK in cell lines of disparate tissue origin; and (iv) luciferase reporter assays showing that PBK promoter activity is dependent on factor binding at intact E2F and CREB/ATF sites. These findings may provide insight into the mechanisms that upregulate PBK expression in proliferative hematologic malignancies and down regulate its expression following growth arrest of leukemic cells.
Nandi
Asit K
AK
University of Maryland School of Medicine, Greenebaum Cancer Center, 655 W Baltimore Street, Baltimore, MD 21201, USA. anandi@som.umaryland.edu
Rapoport
Aaron P
AP
D051696:Activating Transcription Factors / Q000378:metabolism*; D001483:Base Sequence; D001798:Blood Proteins / Q000378:metabolism*; D015153:Blotting, Western; D002453:Cell Cycle; D002460:Cell Line; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D017931:DNA Primers; D050684:E2F Transcription Factors / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D005434:Flow Cytometry; D006801:Humans; D020929:Mitogen-Activated Protein Kinase Kinases; D016297:Mutagenesis, Site-Directed; D011401:Promoter Regions, Genetic; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
D051696:Activating Transcription Factors; D001798:Blood Proteins; D017362:Cyclic AMP Response Element-Binding Protein; D017931:DNA Primers; D050684:E2F Transcription Factors; D012333:RNA, Messenger; D017346:Protein Serine-Threonine Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; C409022:PDZ-binding kinase