Biophysical Characterization of the BIRD Complex and their Mode of Interaction

Abstract

In Arabidopsis thaliana, the development and the defense system are precisely controlled by some proteins to allocate energy and resources as needed. JASMONATE-ZIM domain 3 protein is the repressor of the jasmonic acid defense pathway. JACKDAW (JKD), SHORTSHOOT (SHR), and SCARECROW (SCR) bind together to form the BIRD complex, which regulates root patterning. The transcription factor Teosinte branched1/Cycloidea/Proliferating cell factor 14 (TCP14) also regulates plant development. Recent data shows that JAZ3 and TCP14 interact with JKD and may form a ternary complex, which reveals the study of the five proteins mentioned above may help to understand how defense signals are interpreted during plant growth. The interactions of these five proteins provide a theoretical base to maximize plant fitness and increase crop yield. Using protein purification, microscale thermophoresis, isothermal titration calorimetry, negative staining, X-ray crystallography in this project, we identified JKD interacted with JAZ3, and JKD interacted with TCP14, but they could not form a ternary complex in vitro; SHR/SCR interacted with JAZ3. Those binding results suggests TCP14 and SHR/SCR may have very similar binding site of JKD, and JAZ3 may guide the degradation of the BIRD complex. In structural studies, we resolved the 2D class average that showed the outline of the BIRD complex and it potentially helped to identify how JKD bound to DNA. We also determined the crystal structure of the TCP14 domain, which was an intertwined dimer that possibly uses arginine residues in the N terminus to interact with DNA. These interaction and structure studies of the five proteins provide the basis to understand how defense signals affect plant development.