Analysis and identification of regulatory elements in the promoters of circadian-expressed genes. (a) Frequency of the evening element (EE) and CCA1-binding site (CBS) motifs in the promoters of circadian-regulated genes classified by phase of peak expression. Asterisks indicate phases during which the frequency of promoters containing the motif is significantly different from that of all circadian promoters. Asterisks are placed above the data point to indicate over-representation of the motif and below to indicate under-representation. Both the EE and the CBS are under-represented in promoters of genes with peak expression at circadian time 16. The horizontal lines indicate frequency of the motifs (solid line = EE; dashed line = CBS) in the promoters of all circadian-regulated genes. (b) Tree of putative circadian clock regulatory elements (CCREs) clustered based on sequence similarity is plotted adjacent to a heat map that represents the frequency of each motif in phase-specific subsets of the promoters of genes determined to be circadian regulated in the original analyses of the Covington (left half of heat map) and Edwards (right half of heat map) datasets [6, 7]. In the heat map, dark and light shading represent high and low frequency, respectively. (c-f) Consensus sequences depicted as sequence logos are shown for select clades. (g-j) The phase-specific frequencies of the consensus sequences are plotted in a similar manner as in panel a, except that frequency data are shown for both the Covington (first 24 hours) and Edwards (second 24 hours) datasets and is normalized to the frequency of the sequence in the promoters of all circadian genes. The mean phase-specific frequencies for all the motifs in a clade are shown as dashed lines. For panels a and g to j, white and gray shading represent subjective day and night, respectively.