Regulatory regions of the C elegans genome contain more low-affinity REF-1 transcription-factor binding sites than high-affinity sites

Abstract

Regulatory regions that control transcription generally contain multiple binding sites for sequence-specific, DNA-binding proteins. Such regions are characterized by clusters of cis-acting sites that are more evolutionarily conserved than neighboring nonfunctional sequences. Here, we present evidence that the regulatory regions of the Caenorhabditis elegans genome contain more low-affinity binding sites for REF-1, a Hairy/Enhancer of split family transcription factor that contains two basic Helix-Loop-Helix DNA-binding domains, than high affinity ones. We modified the systematic evolution of ligands by exponential enrichment-serial analysis of gene expression (SELEX-SAGE) procedure to obtain detailed information on the low- and high-affinity DNA-binding sequences for REF-1. We then investigated whether the REF-1 binding sites are involved in any characteristic features of the C elegans genome. Statistics clearly showed that the actual frequencies of both low- and high-affinity binding sites within the genome are significantly higher than the expected frequencies. Computational analysis also shows that both low- and high-affinity binding sites for REF-1 are enriched within the regions of the genome that contain more regulatory motifs, suggesting that both types of sites are involved in regulatory events. The low-affinity sites are enriched more than the high-affinity sites within the evolutionarily conserved regions of the genome, which suggests the possibility that a single high-affinity site can compose a regulatory element, but low-affinity sites must be clustered to become an active regulatory element.