Abstract
BackgroundTranscription factors activate their target genes by binding to specific response elements. Many transcription factor families evolved from a common ancestor by gene duplication and subsequent divergent evolution. Members of the p53 family, which play key roles in cell-cycle control and development, share conserved DNA binding and oligomerisation domains but exhibit distinct functions. In this study, the molecular basis of the functional divergence of related transcription factors was investigated.ResultsWe characterised the DNA-binding specificity and oligomerisation properties of human p53, p63 and p73, as well as p53 from other organisms using novel biophysical approaches. All p53 family members bound DNA cooperatively as tetramers with high affinity. Despite structural differences in the oligomerisation domain, the dissociation constants of the tetramers was in the low nanomolar range for all family members, indicating that the strength of tetramerisation was evolutionarily conserved. However, small differences in the oligomerisation properties were observed, which may play a regulatory role. Intriguingly, the DNA-binding specificity of p53 family members was highly conserved even for evolutionarily distant species. Additionally, DNA recognition was only weakly affected by CpG methylation. Prediction of p53/p63/p73 binding sites in the genome showed almost complete overlap between the different homologs.ConclusionDiversity of biological function of p53 family members is not reflected in differences in sequence-specific DNA binding. Hence, additional specificity factors must exist, which allowed the acquisition of novel functions during evolution while preserving original roles.
Highlights
Transcription factors activate their target genes by binding to specific response elements
Oligomerisation equilibria We have shown previously that full-length human p53 dissociates into dimers at nanomolar concentration, and that oligomerisation is essential for high-affinity DNA binding [21,22]
We used sedimentation velocity analytical ultracentrifugation (SV-AUC) experiments with a fluorescence detection system [23], which allows measurements to be made at low nanomolar concentrations
Summary
Transcription factors activate their target genes by binding to specific response elements. Members of the p53 family, which play key roles in cell-cycle control and development, share conserved DNA binding and oligomerisation domains but exhibit distinct functions. Divergence of transcription factor function within a family could originate from evolutionary changes in the DNA-binding specificity and in the oligomerisation properties. P53 is at the centre of a tumour suppressor network [4,5], and, as such, is essential for the prevention of cancer [6,7] Both p63 and p73 are involved in developmental processes. Each protein has different isoforms [19], which, in most cases, have identical DNA-binding domains but exhibit differences in transcriptional activity, adding an additional layer of complexity [17]
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