Molecular insights into DNA recognition by HD-Zip transcription factors.

Homeodomain-leucine zipper (HD-Zip) genes encode a large family of plant-specific transcription factors (TFs) that are integral to plant development, growth, regulation, and responses to environmental and hormonal signals. While the roles and mechanisms of HD-Zip TFs have been extensively studied, the structural basis for their DNA recognition remains unclear. In this study, we analyzed DAP-seq data and identified consensus DNA motifs, 5'-AAT[W]AT-3' and 5'-[N]AAA[N][N]-3', preferentially bound by HD-Zip TFs. Both motifs feature a 5'-AA(T/A)-3' core, which is shared across previously identified HD-Zip target sequences, suggesting a common recognition feature within the HD-Zip family. Focusing on the well-characterized HD-Zip IV TF PROTEIN PRODUCTION FACTOR 2 (PDF2) from Arabidopsis (Arabidopsis thaliana) and its interaction with the L1 box DNA sequence, our structural and biochemical analyses revealed that the PDF2 HD-ZA module forms a dimer to specifically recognize the 5'-AATG-3' core through an asymmetric binding mode. In this mode, only the primary recognition helix of 1 protomer and the N-arm of the other protomer in the PDF2 HD-ZA dimer are involved in specific DNA interactions. Our study offers insights into the molecular mechanisms of HD-Zip TFs and provides a structural template for engineering applications in agricultural research.