Characterization of MADS-domain transcription factor complexes in Arabidopsis flower development

Abstract

Floral organs are specified by the combinatorial action of MADSdomain transcription factors, yet the mechanisms by which MADSdomain proteins activate or repress the expression of their target genes and the nature of their cofactors are still largely unknown. Here, we show using affinity purification and mass spectrometry that five major floral homeotic MADS-domain proteins (AP1, AP3, PI, AG, and SEP3) interact in floral tissues as proposed in the "floral quartet" model. In vitro studies confirmed a flexible composition of MADSdomain protein complexes depending on relative protein concentrations and DNA sequence. In situ bimolecular fluorescent complementation assays demonstrate that MADS-domain proteins interact during meristematic stages of flower development. By applying a targeted proteomics approach we were able to establish a MADS-domain protein interactome that strongly supports a mechanistic link between MADS-domain proteins and chromatin remodeling factors. Furthermore, members of other transcription factor families were identified as interaction partners of floral MADS-domain proteins suggesting various specific combinatorial modes of action.protein complex isolation | transcriptional regulation | chromatin activation | histone marks F lower development is one of the best understood developmental processes in plants. According to the classic ABC model (1), floral organs in the model plant species Arabidopsis are specified by the combinatorial activity of three functional gene classes. The A class genes represented by APETALA1 (AP1) and APETALA2 (AP2) specify sepal identity, and together with B class genes APETALA3 (AP3) and PISTILLATA (PI), they determine the identity of petals. The C class gene AGA-MOUS (AG) alone determines carpel identity and, together with B class genes, it specifies stamen identity. The ABC model was extended to the ABCE model, in which E class genes [SEPAL-LATA1-4 (SEP1-4)] are required for the specification of all four types of floral organs (2, 3). Based on genetic and yeast n-hybrid protein interaction data, it was later proposed in the "floral quartet model" that floral organs are specified by combinatorial protein interactions of ABCE-class MADS-domain transcription factors, which are thought to assemble into organ-specific quaternary protein complexes that bind to two CArG boxes, DNA consensus sequence CC[A/T] 6 GG, in regulatory regions of target genes (4, 5). E-class proteins have a special role in this model as major mediators of higher-order complex formation. Although interactions that were predicted in this model were further supported by additional in vitro DNA-binding assays and protoplast , formation and composition of these complexes in endogenous tissues remained unknown.Heterologous interaction studies in yeast and genetic data suggest recruitment of transcriptional coregulators such as SEUSS (SEU) and LEUNIG (LUG) by floral MADS-domain proteins (9). Ovule-specific MADS-domain protein complexes were found to form higher-order interactions with BELL1 (BEL1), a mem...

SWP73 Subunits of Arabidopsis SWI/SNF Chromatin Remodeling Complexes Play Distinct Roles in Leaf and Flower Development

“…The SEP1-4 genes are required for the specification of all four types of floral organs. Expression of SEP3 is directly regulated by AP1 and several other MADS box proteins (Smaczniak et al, 2012). In comparison, the swp73a mutation had no significant effect on the expression of these floral organ identity genes.…”

“…Furthermore, the swp73b-1 mutant exhibited nucleosome stabilization and altered nucleosome occupancy around position 2670 of the AP1 locus, just upstream of known binding sites of transcription factors LFY, LMI2, SPL, and FT/FD (Pastore et al, 2011). In addition to altered nucleosome positioning in the region 2740 to 21120, fuzzy nucleosome arrangement was suggested by the MNase-Seq data in swp73b-1 mutant between positions 23000 and 22800 of SEP3, which carries the entry site of a transcription regulatory complex of MADS box proteins SEP3, AP1, AG, and FUL (Smaczniak et al, 2012). Nucleosome stabilization was detected around position 2270 of the AP3 promoter corresponding to a CArG-box motif (Tilly et al, 1998), which is also known to bind the BRM and SPLAYED (SYD) ATPase subunits of SWI/SNF complexes .…”

RiceLHS1/OsMADS1Controls Floret Meristem Specification by Coordinated Regulation of Transcription Factors and Hormone Signaling Pathways

“…cytokinin pathway components) its target genes. We speculate that its interactions with partners in higher order complexes, and their interactions with chromatin modifiers, may determine its effect on gene expression, as is known for other transcription factor complexes (Sridhar et al, 2006;Liu et al, 2007;Smaczniak et al, 2012). The cumulative effects of these complexes on net target gene expression status culminates in a rice spikelet with a determinate floret with differentiated organs (Fig.…”

Dynamic regulation of Polycomb group activity during plant development

“…Although the function of PRC2 in the regulation of cell proliferation and differentiation is conserved throughout plants [58], fine-tuning is likely achieved through the various mechanisms described above, which may be lineage-specific. The recent development of an efficient targeted technique to isolate protein complexes in plants may accelerate the identification of PRC2-associated co-factors in the future [59]. Temporal and spatial control of PRC2 repression is achieved through association of the core PRC2 complex with cell-type specific co-factors.…”