Identification and functional characterization of the human and murine polo-like kinase (Plk) promoter.

The Plk gene encodes a serine/theronine kinase which is located in the nucleus. Northern blot analysis linked Plk expression to the proliferative activity of cells and tissues. To analyse the transcriptional regulation of the Plk gene we have isolated several human genomic clones containing the Plk promoter. RNAse protection assays revealed three major transcription start sites within a 40 bp region centered around the 5' end of the known human cDNA and 6 minor Cap sites. A genomic fragment of 2.3 kb located 5' to the translation start sites drives the expression of the CAT-reporter in transient transfections in human (EPLC, HeLa) and mouse (NIH3T3, 32D) cell lines in an orientation dependent fashion. The 2.3 kb genomic fragment contains a CCAAT motif located 30-70 bp upstream of the Cap sites and two overlapping Sp1 sites 20 bp further upstream. Additional sequence motif homologues to binding sites of known transcription factors could be identified. In addition to the human Plk promoter, the mouse Plk promoter was isolated. The sequence alignment of the human and murine promoter revealed three regions with extensive sequence homology within a region of 300 bp immediately upstream of the Cap sites. A fourth region of homology encompassing 90 bp about 2.1 kb 5' of the Cap sites was identified as well. Deletion of various regions within the 2.3 kb promoter fragment identified several domains involved in the regulation of the human Plk promoter. The 300 bp region immediately 5' of the Cap sites which is highly conserved between mouse and man is essential for promoter activity. 3' deletions including the CCAAT site abolished promoter activity. Growing 5' deletions within the core region of the promoter reduces transcriptional activity. Furthermore, using deletion clones we identified regions 5' of the core region which enhance or silence the transcriptional activity of the core promoter.