Structural Basis for the Oligomerization of the MADS Domain Transcription Factor SEPALLATA3 in Arabidopsis  

Abstract

In plants, MADS domain transcription factors act as central regulators of diverse developmental pathways. In Arabidopsis thaliana, one of the most central members of this family is SEPALLATA3 (SEP3), which is involved in many aspects of plant reproduction, including floral meristem and floral organ development. SEP3 has been shown to form homo and heterooligomeric complexes with other MADS domain transcription factors through its intervening (I) and keratin-like (K) domains. SEP3 function depends on its ability to form specific protein-protein complexes; however, the atomic level determinants of oligomerization are poorly understood. Here, we report the 2.5-Å crystal structure of a small portion of the intervening and the complete keratin-like domain of SEP3. The domains form two amphipathic alpha helices separated by a rigid kink, which prevents intramolecular association and presents separate dimerization and tetramerization interfaces comprising predominantly hydrophobic patches. Mutations to the tetramerization interface demonstrate the importance of highly conserved hydrophobic residues for tetramer stability. Atomic force microscopy was used to show SEP3-DNA interactions and the role of oligomerization in DNA binding and conformation. Based on these data, the oligomerization patterns of the larger family of MADS domain transcription factors can be predicted and manipulated based on the primary sequence.

MADS-domain transcription factors and the floral quartet model of flower development: linking plant development and evolution

“…Recent X-ray crystallography studies of the K domain of A. thaliana SEP3 revealed that the K domain forms two amphipathic α-helices separated by a rigid kink, which prevents intramolecular association (Puranik et al, 2014). The K domain thus provides two separate interaction interfaces to facilitate dimerization and tetramerization with other K domains (Puranik et al, 2014). Atomic force microscopy (AFM) further demonstrated the looping of target DNA by SEP3 and even allowed FQCs to be 'seen' for the first time (Puranik et al, 2014).…”

“…The K domain thus provides two separate interaction interfaces to facilitate dimerization and tetramerization with other K domains (Puranik et al, 2014). Atomic force microscopy (AFM) further demonstrated the looping of target DNA by SEP3 and even allowed FQCs to be 'seen' for the first time (Puranik et al, 2014).…”

“…Some EMSA experiments had demonstrated that the C-terminal half of the K domain, which was assumed to form an amphipathic α-helix involved in the formation of a coiled-coil, is of crucial importance for MIKC-type protein tetramerization ). Recent X-ray crystallography studies of the K domain of A. thaliana SEP3 revealed that the K domain forms two amphipathic α-helices separated by a rigid kink, which prevents intramolecular association (Puranik et al, 2014). The K domain thus provides two separate interaction interfaces to facilitate dimerization and tetramerization with other K domains (Puranik et al, 2014).…”

“…Among Given that the formation of functional tetramers is a widespread property of A. thaliana MIKC-type proteins (Puranik et al, 2014;Espinosa-Soto et al, 2014), we hypothesize that FQCs play important roles beyond floral organ identity specification. Indeed, several studies have suggested that MIKC-type proteins other than the canonical class A-E floral homeotic proteins of the FQM can form FQCs.…”

“…The I domain, by contrast, is only relatively weakly conserved and contributes to the selective formation of DNA-binding dimers (Kaufmann et al, 2005b). The K domain is characterized by a conserved, regular spacing of hydrophobic and charged residues, which allows the formation of amphipathic helices involved in protein dimerization and multimeric complex formation (Yang et al, 2003;Puranik et al, 2014). Finally, the C domain is quite variable and, in some MADSdomain proteins, is involved in transcriptional activation or multimeric complex formation (for a review on structural and phylogenetic aspects of MIKC-type proteins, see Kaufmann et al, 2005b;Theißen and Gramzow, 2016).…”