Dark Matter of the Transcription Factor Binding Site Motif Universe

Background: We study the limits imposed by transcription factor specificity on the maximum size of a genetic regulatory network. Results: Most regular expressions for natural transcription factor binding site motifs are separated in sequence space by only one to three motif-discriminating positions. This mild specificity requirement puts the number of transcription factors that can coexist with minimal crosstalk on the order of ten thousand, which would fully utilize the space of DNA subsequences. An expanded alphabet with modified bases can further raise this limit by several orders of magnitude, at the expense of sequence space usage. Conclusions: Based on this analysis, thousands of transcription factor binding site motifs may await discovery.