Gene alterations that affect transcription control networks between species have the potential to modify relative gene expression profile signatures or even disease model processes, for example mouse models of human cancer. Transcriptional networks can trans-evolve through variation of master regulators, and/or cis-evolve by changes in the regulation of target genes that can themselves play roles in networks. The tumor suppressor p53 is a sequence-specific transcription factor that can be considered a master regulator of stress responses. The mouse and human p53 proteins are ∼90% identical in their DNA binding domains and exhibited identical transactivation capacities towards 20 response elements (REs) (Resnick and Inga, PNAS, 2003). In order to address the evolution of p53 networks due to variation in RE sequences we have chosen to examine the conservation in mice of well-established p53 REs identified in human p53 target genes. Since the consensus p53 RE is degenerate (two copies of 5′-RRRC(A/T)(T/A)GYYY-3′) and most of the established p53 REs contain mismatches, we have addressed both conservation of RE sequence and functionality in p53-dependent transactivation. We developed a customized bioinformatics approach combining phylogenetic footprint and a set of response element rules, derived from yeast-based functional assays, that describe the transactivation potential of defined RE sequences. We examined 26 established p53 target genes belonging to different functional groupings including cell cycle control, apoptosis, DNA repair and angiogenesis. First, we asked whether the established p53 target REs in humans are conserved in the homologous promoter region in mouse. We then searched the flanking (5′ and 3′) 5 kb regions of the transcriptional start site (TSS) in both species for the incidence of additional p53 functional REs. Furthermore, for the case of mouse and human RE sequence divergence, yeast and mammalian assays were used to compare p53 transactivation towards the isolated REs as well as large promoter regions from both species. We found that ∼60% of the REs are either functionally conserved or at least have the ability to respond to p53 from REs located at different positions in the promoters of the two species. Surprisingly, most of the genes involved in DNA metabolism and repair showed no conservation of RE sequence nor compensatory REs in the 10 kb TSS flanking regions. Our results with p53 indicate that a true assessment of evolution of transcriptional regulatory networks using computational approaches should include the analysis of functional conservation based on response element rules that are established in vivo. The differences in p53-mediated modulation of DNA metabolism in humans and mice may contribute to differences in stress responses as well as cancer development between the species. A Inga, D. Menendez and A. Jegga contributed equally to the work.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]