Many transcription factor families have evolutionarily conserved binding motifs in plants.

Transcription factors control gene expression during development and in response to a broad range of internal and external stimuli. They regulate promoter activity by directly binding cis-regulatory elements in DNA. The angiosperm Arabidopsis (Arabidopsis thaliana) contains more than 1,500 annotated transcription factors, each containing a DNA-binding domain that is used to define transcription factor families. Analyzing the binding motifs of 686 and the binding sites of 335 Arabidopsis transcription factors, as well as motifs of 92 transcription factors from other plants, we identified a constrained vocabulary of 74 conserved motifs spanning 50 families in plants. Among 21 transcription factor families, we found 1 core motif for all analyzed members and between 2% and 72% overlapping binding sites. Five families show conservation of the motif along phylogenetic clades. Five families, including the C2H2 zinc finger family, show high diversity among motifs in plants, suggesting potential for the neofunctionalization of duplicated transcription factors based on the motif recognized. We tested whether conserved motifs remained conserved since at least 450 million years ago by determining the binding motifs of 17 transcription factors from 11 families in Marchantia (Marchantia polymorpha) using amplified DNA affinity purification sequencing. We detected nearly identical binding motifs as predicted from the angiosperm data. Our findings show a large repertoire of overlapping binding sites within a transcription factor family and species and a high degree of binding motif conservation for at least 450 million years, indicating more potential for evolution in cis- rather than trans-regulatory elements.