Brendan Davies

Molecular and Phylogenetic Analyses of the Complete MADS-Box Transcription Factor Family in Arabidopsis: New Openings to the MADS World

MADS-box transcription factors are key regulators of several plant development processes. Analysis of the complete Arabidopsis genome sequence revealed 107 genes encoding MADS-box proteins, of which 84% are of unknown function. Here, we provide a complete overview of this family, describing the gene structure, gene expression, genome localization, protein motif organization, and phylogenetic relationship of each member. We have divided this transcription factor family into five groups (named MIKC, Mα, Mβ, Mγ, and Mδ) based on the phylogenetic relationships of the conserved MADS-box domain. This study provides a solid base for functional genomics studies into this important family of plant regulatory genes, including the poorly characterized group of M-type MADS-box proteins. MADS-box genes also constitute an excellent system with which to study the evolution of complex gene families in higher plants.

Comprehensive Interaction Map of the Arabidopsis MADS Box Transcription Factors

Interactions between proteins are essential for their functioning and the biological processes they control. The elucidation of interaction maps based on yeast studies is a first step toward the understanding of molecular networks and provides a framework of proteins that possess the capacity and specificity to interact. Here, we present a comprehensive plant protein–protein interactome map of nearly all members of the Arabidopsis thaliana MADS box transcription factor family. A matrix-based yeast two-hybrid screen of >100 members of this family revealed a collection of specific heterodimers and a few homodimers. Clustering of proteins with similar interaction patterns pinpoints proteins involved in the same developmental program and provides valuable information about the participation of uncharacterized proteins in these programs. Furthermore, a model is proposed that integrates the floral induction and floral organ formation networks based on the interactions between the proteins involved. Heterodimers between flower induction and floral organ identity proteins were observed, which point to (auto)regulatory mechanisms that prevent the activity of flower induction proteins in the flower.

PLENA and FARINELLI: redundancy and regulatory interactions between two Antirrhinum MADS‐box factors controlling flower development

We report the discovery of an Antirrhinum MADS‐box gene, FARINELLI (FAR), and the isolation of far mutants by a reverse genetic screen. Despite striking similarities between FAR and the class C MADS‐box gene PLENA (PLE), the phenotypes of their respective mutants are dramatically different. Unlike ple mutants, which show homeotic conversion of reproductive organs to perianth organs and a loss of floral determinacy, far mutants have normal flowers which are partially male‐sterile. Expression studies of PLE and FAR, in wild‐type and mutant backgrounds, show complex interactions between the two genes. Double mutant analysis reveals an unexpected, redundant negative control over the B‐function MADS‐box genes. This feature of the two Antirrhinum C‐function‐like genes is markedly different from the control of the inner boundary of the B‐function expression domain in Arabidopsis, and we propose and discuss a model to account for these differences. The difference in phenotypes of mutants in two highly related genes illustrates the importance of the position within the regulatory network in determining gene function.

The TOPLESS Interactome: A Framework for Gene Repression in Arabidopsis

Transcription factors activate or repress target gene expression or switch between activation and repression. In animals and yeast, Groucho/Tup1 corepressor proteins are recruited by diverse transcription factors to induce context-specific transcriptional repression. Two groups of Groucho/Tup1-like corepressors have been described in plants. LEUNIG and LEUNIG_HOMOLOG constitute one group and TOPLESS (TPL) and the four TPL-related (TPR) corepressors form the other. To discover the processes in which TPL and the TPR corepressors operate, high-throughput yeast two-hybrid approaches were used to identify interacting proteins. We found that TPL/TPR corepressors predominantly interact directly with specific transcription factors, many of which were previously implicated in transcriptional repression. The interacting transcription factors reveal that the TPL/TPR family has been coopted multiple times to modulate gene expression in diverse processes, including hormone signaling, stress responses, and the control of flowering time, for which we also show biological validation. The interaction data suggest novel mechanisms for the involvement of TPL/TPR corepressors in auxin and jasmonic acid signaling. A number of short repression domain (RD) sequences have previously been identified in Arabidopsis (Arabidopsis thaliana) transcription factors. All known RD sequences were enriched among the TPL/TPR interactors, and novel TPL-RD interactions were identified. We show that the presence of RD sequences is essential for TPL/TPR recruitment. These data provide a framework for TPL/TPR-dependent transcriptional repression. They allow for predictions about new repressive transcription factors, corepressor interactions, and repression mechanisms and identify a wide range of plant processes that utilize TPL/TPR-mediated gene repression.

Floral organ identity: 20 years of ABCs

One of the early successes of the application of molecular genetics to study plant development was the discovery of a series of genes that act together, in an apparently simple combinatorial model, to specify the identity of the different organs of a flower. Widely known as the ABC model, this framework for understanding has been investigated and modified over the course of the last two decades. The cast list of genes has been defined and, as other chapters in this volume will show, great progress has been made in understanding how they are regulated, how they act together, what they do and how they have contributed to the evolution of the flower in its varied forms. In this introductory review to the volume we will review the derivation and elaboration of the most current version of the ABC model, highlighting the modifications that have been necessary to ensure it fits the available experimental data. We will highlight the remaining difficulties in fitting the current model to the experimental data and propose a further modification to enable it to regain its applicability.

Analysis of the Transcription Factor WUSCHEL and Its Functional Homologue in Antirrhinum Reveals a Potential Mechanism for Their Roles in Meristem Maintenance

One of the most significant features of plant development is the way in which it can be elaborated and modulated throughout the life of the plant, an ability that is conferred by meristems. The Arabidopsis thaliana WUSCHEL gene (WUS), which encodes a homeodomain transcription factor, is required to maintain the stem cells in the shoot apical meristem in an undifferentiated state. The mechanism by which WUS prevents the differentiation of stem cells is unknown. We have characterized a meristem maintenance mutant in Antirrhinum majus and shown that it arises from a defect in the WUS orthologue ROSULATA (ROA). Detailed characterization of a semidominant roa allele revealed an essential role for the conserved C-terminal domain. Expression of either ROA or WUS lacking this domain causes a failure of meristem maintenance. The conserved domain mediates an interaction between WUS and two members of a small family of corepressor-like proteins in Arabidopsis. Our results suggest that WUS functions by recruiting transcriptional corepressors to repress target genes that promote differentiation, thereby ensuring stem cell maintenance.

Evolution in Action: Following Function in Duplicated Floral Homeotic Genes

Gene duplication plays a fundamental role in evolution by providing the genetic material from which novel functions can arise. Newly duplicated genes can be maintained by subfunctionalization (the duplicated genes perform different aspects of the original genes function) and/or neofunctionalization (one of the genes acquires a novel function). PLENA in Antirrhinum and AGAMOUS in Arabidopsis are the canonical C-function genes that are essential for the specification of reproductive organs. These functionally equivalent genes encode closely related homeotic MADS-box transcription factors. Using genome synteny, we confirm phylogenetic analyses showing that PLENA and AGAMOUS are nonorthologous genes derived from a duplication in a common ancestor. Their respective orthologs, SHATTERPROOF in Arabidopsis and FARINELLI in Antirrhinum, have undergone independent subfunctionalization via changes in regulation and protein function. Surprisingly, the functional divergence between PLENA and FARINELLI, is morphologically manifest in both transgenic Antirrhinum and Arabidopsis. This provides a clear illustration of a random evolutionary trajectory for gene functions after a duplication event. Different members of a duplicated gene pair have retained the primary homeotic functions in different lineages, illustrating the role of chance in evolution. The differential ability of the Antirrhinum genes to promote male or female development provides a striking example of subfunctionalization at the protein level.

CUPULIFORMIS establishes lateral organ boundaries in Antirrhinum

cupuliformis mutants are defective in shoot apical meristem formation, but cup plants overcome this early barrier to development to reach maturity. CUP encodes a NAC-domain transcription factor, homologous to the Petunia NAM and Arabidopsis CUC proteins. The phenotype of cup mutants differs from those of nam and cuc1 cuc2 in that dramatic organ fusion is observed throughout development. In addition to cotyledon and floral organ fusions, severe lateral organ fusion is found in leaves and inflorescences, and the apical meristem becomes highly fasciated. These features reveal a role for CUP in the establishment of all above ground organ boundaries. Consistent with this function, CUP is expressed at the boundaries of all lateral organs and meristems. It is not currently known how NAC-domain genes act to establish organ boundaries. Here, we show that CUP directly interacts with a TCP-domain transcription factor. Members of the TCP-domain family have previously been shown to regulate organ outgrowth. Our results suggest a model for the establishment of organ boundaries based on the localised expression of NAC-domain and TCP-domain factors.

Formins: intermediates in signal-transduction cascades that affect cytoskeletal reorganization

The control of cell growth and polarity depends on a dynamic actin cytoskeleton that has the ability to reorganize in response to developmental and environmental stimuli. In animals and fungi, formins are just one of the four major classes of poly-L-proline-containing (PLP) proteins that form part of the signal-transduction cascade that leads to rearrangement of the actin cytoskeleton. Analysis of the Arabidopsis genome sequence indicates that, unlike animals and fungi, formins are the only class of conserved profilin-binding PLP proteins in plants. Moreover, plant formins show significant structural differences compared with their animal and fungal counterparts, raising the possibility that plant formins are subject to novel mechanisms of control or perform unique roles in plants.

TCP14 and TCP15 affect internode length and leaf shape in Arabidopsis

TCP transcription factors constitute a small family of plant-specific bHLH-containing, DNA-binding proteins that have been implicated in the control of cell proliferation in plants. Despite the significant role that is likely to be played by genes that control cell division in the elaboration of plant architecture, functional analysis of this family by forward and reverse genetics has been hampered by genetic redundancy. Here we show that mutants in two related class I TCP genes display a range of growth-related phenotypes, consistent with their dynamic expression patterns; these phenotypes are enhanced in the double mutant. Together, the two genes influence plant stature by promoting cell division in young internodes. Reporter gene analysis and use of SRDX fusions suggested that TCP14 and TCP15 modulate cell proliferation in the developing leaf blade and specific floral tissues; a role that was not apparent in our phenotypic analysis of single or double mutants. However, when the relevant mutants were subjected to computer-aided morphological analysis of the leaves, the consequences of loss of either or both genes became obvious. The effects on cell proliferation of perturbing the function of TCP14 and TCP15 vary with tissue, as has been suggested for other TCP factors. These findings indicate that the precise elaboration of plant form is dependent on the cumulative influence of many TCP factors acting in a context-dependent fashion. The study highlights the need for advanced methods of phenotypic analysis in order to characterize phenotypes and to construct a dynamic model for TCP gene function.