A conserved leucine zipper-like motif accounts for strong tetramerization capabilities of SEPALLATA-like MADS-domain transcription factors.

Florian Rümpler,Rainer Melzer,Günter Theißen

doi:10.1093/jxb/ery063

Abstract

The development of angiosperm flowers is regulated by homeotic MIKC-type MADS-domain transcription factors that activate or repress target genes via the formation of DNA-bound, organ-specific tetrameric complexes. The protein-protein interaction (PPI) capabilities differ considerably between different MIKC-type proteins. In Arabidopsis thaliana the floral homeotic protein SEPALLATA3 (SEP3) acts as a hub that incorporates numerous other MADS-domain proteins into tetrameric complexes that would otherwise not form. However, the molecular mechanisms that underlie these promiscuous interactions remain largely unknown. In this study, we created a collection of amino acid substitution mutants of SEP3 to quantify the contribution of individual residues on protein tetramerization during DNA-binding, employing methods of molecular biophysics. We show that leucine residues at certain key positions form a leucine-zipper structure that is essential for tetramerization of SEP3, whereas the introduction of physicochemically very similar residues at respective sites impedes the formation of DNA-bound tetramers. Comprehensive molecular evolutionary analyses of MADS-domain proteins from a diverse set of flowering plants revealed exceedingly high conservation of the identified leucine residues within SEP3-subfamily proteins throughout angiosperm evolution. In contrast, MADS-domain proteins that are unable to tetramerize among themselves exhibit preferences for other amino acids at homologous sites. Our findings indicate that the subfamily-specific conservation of amino acid residues at just a few key positions accounts for subfamily-specific interaction capabilities of MADS-domain transcription factors and this has shaped the present-day structure of the PPI network controlling flower development.

Highlights

Complexity of biological systems is often achieved by the combined activity of a small number of factors (Reményi et al, 2004)
One important example is represented by protein–protein interaction (PPI) networks that are based on transcription factors (TFs) that act in a combinatorial manner to accomplish the required degree of (e.g.) morphological complexity
Beyond the three K-subdomains, we introduced proline substitutions at positions occupied by two conserved hydrophobic amino acids in the interhelical region between the K1- and the K2-subdomain (L120P and L123P, Fig. 2A) because L120 and L123 are homologous to L121 and V124 in the MADS-domain protein PI and those positions have been shown to be important for protein–protein interactions (Yang and Jack, 2004)

Summary

Introduction

Complexity of biological systems is often achieved by the combined activity of a small number of factors (Reményi et al, 2004). One important example is represented by protein–protein interaction (PPI) networks that are based on transcription factors (TFs) that act in a combinatorial manner to accomplish the required degree of (e.g.) morphological complexity. Importance for almost all developmental processes, the molecular determinants that underlie the specific combinatorial interactions remain poorly understood.This is especially true for PPIs among transcription factors (TFs) belonging to the same family. The four SEP proteins act in a largely redundant manner but, in agreement with their central position in the PPI network controlling flower development, sep multiple-mutants show severe developmental defects (Pelaz et al, 2000; Ditta et al, 2004). The four SEP proteins act in a largely redundant manner but, in agreement with their central position in the PPI network controlling flower development, sep multiple-mutants show severe developmental defects (Pelaz et al, 2000; Ditta et al, 2004). sep sep sep triple-mutant plants develop sepals from primordia that would normally develop into petals, stamens, and carpels, and sep sep sep sep quadruplemutants develop vegetative leaves instead of floral organs (Pelaz et al, 2000; Ditta et al, 2004)

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