Chk1
kinase is a key signal transducer in the checkpoint pathways responding to DNA
damage. Chk1 has been shown to be transcriptionally down-regulated in
response to cis-dichloro-diamino platinum treatment in a p53-dependent
manner. Now we isolated and characterised the 5' flanking region of the
Chk1 gene. We demonstrated that the isolated region has promoter activity
when cloned upstream to a reporter luciferase gene and we could define the
minimal promoter region. The modulation of the transcriptional activity of
the cloned Chk1 promoter region by different transcription factors was investigated
by co-transfection experiments and by using different isogenic systems. It
was shown that p53 is indeed able to down-regulate the promoter activity of
the cloned region, providing a mechanistic explanation to the observation
that p53 decreases Chk1 protein levels after DNA damage. Several E2F
binding sites were detected in the genomic sequence and E2F1 induced Chk1
promoter activity in co-transfection experiments. This induction was
abolished when a mutated form of E2F1, not able to bind DNA, was used.
Over-expression of E2F1 sustained an increased Chk1 promoter activity,
whereas overexpression of Rb protein, which binds E2F factors, decreased
Chk1 promoter activity. We discuss our results in the context of Chk1
involvment in DNA damage checkpoint pathways.
