Cauliflower fractal forms arise from perturbations of floral gene networks.

Throughout development, plant meristems regularly produce organs in defined spiral, opposite, or whorl patterns. Cauliflowers present an unusual organ arrangement with a multitude of spirals nested over a wide range of scales. How such a fractal, self-similar organization emerges from developmental mechanisms has remained elusive. Combining experimental analyses in an Arabidopsis thaliana cauliflower-like mutant with modeling, we found that curd self-similarity arises because the meristems fail to form flowers but keep the "memory" of their transient passage in a floral state. Additional mutations affecting meristem growth can induce the production of conical structures reminiscent of the conspicuous fractal Romanesco shape. This study reveals how fractal-like forms may emerge from the combination of key, defined perturbations of floral developmental programs and growth dynamics.

Selection and gene flow shape genomic islands that control floral guides.

Genomes of closely-related species or populations often display localized regions of enhanced relative sequence divergence, termed genomic islands. It has been proposed that these islands arise through selective sweeps and/or barriers to gene flow. Here, we genetically dissect a genomic island that controls flower color pattern differences between two subspecies of Antirrhinum majus, A.m.striatum and A.m.pseudomajus, and relate it to clinal variation across a natural hybrid zone. We show that selective sweeps likely raised relative divergence at two tightly-linked MYB-like transcription factors, leading to distinct flower patterns in the two subspecies. The two patterns provide alternate floral guides and create a strong barrier to gene flow where populations come into contact. This barrier affects the selected flower color genes and tightly-linked loci, but does not extend outside of this domain, allowing gene flow to lower relative divergence for the rest of the chromosome. Thus, both selective sweeps and barriers to gene flow play a role in shaping genomic islands: sweeps cause elevation in relative divergence, while heterogeneous gene flow flattens the surrounding "sea," making the island of divergence stand out. By showing how selective sweeps establish alternative adaptive phenotypes that lead to barriers to gene flow, our study sheds light on possible mechanisms leading to reproductive isolation and speciation.

Evolution of flower color pattern through selection on regulatory small RNAs.

Small RNAs (sRNAs) regulate genes in plants and animals. Here, we show that population-wide differences in color patterns in snapdragon flowers are caused by an inverted duplication that generates sRNAs. The complexity and size of the transcripts indicate that the duplication represents an intermediate on the pathway to microRNA evolution. The sRNAs repress a pigment biosynthesis gene, creating a yellow highlight at the site of pollinator entry. The inverted duplication exhibits steep clines in allele frequency in a natural hybrid zone, showing that the allele is under selection. Thus, regulatory interactions of evolutionarily recent sRNAs can be acted upon by selection and contribute to the evolution of phenotypic diversity.

Separate elements of the TERMINAL FLOWER 1 cis-regulatory region integrate pathways to control flowering time and shoot meristem identity.

TERMINAL FLOWER 1 (TFL1) is a key regulator of Arabidopsis plant architecture that responds to developmental and environmental signals to control flowering time and the fate of shoot meristems. TFL1 expression is dynamic, being found in all shoot meristems, but not in floral meristems, with the level and distribution changing throughout development. Using a variety of experimental approaches we have analysed the TFL1 promoter to elucidate its functional structure. TFL1 expression is based on distinct cis-regulatory regions, the most important being located 3' of the coding sequence. Our results indicate that TFL1 expression in the shoot apical versus lateral inflorescence meristems is controlled through distinct cis-regulatory elements, suggesting that different signals control expression in these meristem types. Moreover, we identified a cis-regulatory region necessary for TFL1 expression in the vegetative shoot and required for a wild-type flowering time, supporting that TFL1 expression in the vegetative meristem controls flowering time. Our study provides a model for the functional organisation of TFL1 cis-regulatory regions, contributing to our understanding of how developmental pathways are integrated at the genomic level of a key regulator to control plant architecture.

Changing the spatial pattern of TFL1 expression reveals its key role in the shoot meristem in controlling Arabidopsis flowering architecture.

Models for the control of above-ground plant architectures show how meristems can be programmed to be either shoots or flowers. Molecular, genetic, transgenic, and mathematical studies have greatly refined these models, suggesting that the phase of the shoot reflects different genes contributing to its repression of flowering, its vegetativeness ('veg'), before activators promote flower development. Key elements of how the repressor of flowering and shoot meristem gene TFL1 acts have now been tested, by changing its spatiotemporal pattern. It is shown that TFL1 can act outside of its normal expression domain in leaf primordia or floral meristems to repress flower identity. These data show how the timing and spatial pattern of TFL1 expression affect overall plant architecture. This reveals that the underlying pattern of TFL1 interactors is complex and that they may be spatially more widespread than TFL1 itself, which is confined to shoots. However, the data show that while TFL1 and floral genes can both act and compete in the same meristem, it appears that the main shoot meristem is more sensitive to TFL1 rather than floral genes. This spatial analysis therefore reveals how a difference in response helps maintain the 'veg' state of the shoot meristem.

Conserved intragenic elements were critical for the evolution of the floral C-function.

The floral C-function, which specifies stamen and carpel development, played a pivotal role in the evolution of flowers. An important aspect of this was the establishment of mechanisms regulating the temporal and spatial expression domain of the C-function genes. Transcription of the Arabidopsis C-function gene AGAMOUS (AG) is tightly controlled by factors that interact with cis-elements within its large second intron. Little is known about the regulatory role of intragenic elements in C-function genes from species other than Arabidopsis. We show that a binding site for the LEAFY (LFY) transcription factor, present in the AG intron, is conserved in the introns of diverse C-function genes and is positioned close to other conserved motifs. Using an in planta mutagenesis approach, we targeted evolutionarily conserved sequences in the intron of the Antirrhinum PLENA (PLE) gene to establish whether they regulate PLE expression. Small sequence deletions resulted in a novel class of heterochronic C-function mutants with delayed onset of PLE expression and loss of stamen identity. These phenotypes differ significantly from weak C-function mutant alleles in Antirrhinum and Arabidopsis. Our findings demonstrate that the PLE intron contains regulatory cis-elements, including a LFY-binding site, critical for establishing the correct C-function expression domain. We show that the LFY site, and other conserved intron elements, pre-date the divergence of the monocot and dicot lineages, suggesting that they were a determinant in the evolution of the C-function, and propose a threshold model to explain phenotypic divergence observed between C-function mutants.

TERMINAL FLOWER1 is a mobile signal controlling Arabidopsis architecture.

Shoot meristems harbor stem cells that provide key growing points in plants, maintaining themselves and generating all above-ground tissues. Cell-to-cell signaling networks maintain this population, but how are meristem and organ identities controlled? TERMINAL FLOWER1 (TFL1) controls shoot meristem identity throughout the plant life cycle, affecting the number and identity of all above-ground organs generated; tfl1 mutant shoot meristems make fewer leaves, shoots, and flowers and change identity to flowers. We find that TFL1 mRNA is broadly distributed in young axillary shoot meristems but later becomes limited to central regions, yet affects cell fates at a distance. How is this achieved? We reveal that the TFL1 protein is a mobile signal that becomes evenly distributed across the meristem. TFL1 does not enter cells arising from the flanks of the meristem, thus allowing primordia to establish their identity. Surprisingly, TFL1 movement does not appear to occur in mature shoots of leafy (lfy) mutants, which eventually stop proliferating and convert to carpel/floral-like structures. We propose that signals from LFY in floral meristems may feed back to promote TFL1 protein movement in the shoot meristem. This novel feedback signaling mechanism would ensure that shoot meristem identity is maintained and the appropriate inflorescence architecture develops.

Control of cell and petal morphogenesis by R2R3 MYB transcription factors.

Petals of animal-pollinated angiosperms have adapted to attract pollinators. Factors influencing pollinator attention include colour and overall size of flowers. Colour is determined by the nature of the pigments, their environment and by the morphology of the petal epidermal cells. Most angiosperms have conical epidermal cells, which enhance the colour intensity and brightness of petal surfaces. The MYB-related transcription factor MIXTA controls the development of conical epidermal cells in petals of Antirrhinum majus. Another gene encoding an R2R3 MYB factor very closely related to MIXTA, AmMYBML2, is also expressed in flowers of A. majus. We have analysed the roles of AmMYBML2 and two MIXTA-related genes, PhMYB1 from Petunia hybrida and AtMYB16 from Arabidopsis thaliana, in petal development. The structural similarity between these genes, their comparable expression patterns and the similarity of the phenotypes they induce when ectopically expressed in tobacco, suggest they share homologous functions closely related to, but distinct from, that of MIXTA. Detailed phenotypic analysis of a phmyb1 mutant confirmed the role of PhMYB1 in the control of cell morphogenesis in the petal epidermis. The phmyb1 mutant showed that epidermal cell shape affects petal presentation, a phenotypic trait also observed following re-examination of mixta mutants. This suggests that the activity of MIXTA-like genes also contributes to petal form, another important factor influencing pollinator attraction.

A single amino acid converts a repressor to an activator of flowering.

Homologous proteins occurring through gene duplication may give rise to novel functions through mutations affecting protein sequence or expression. Comparison of such homologues allows insight into how morphological traits evolve. However, it is often unclear which changes are key to determining new functions. To address these ideas, we have studied a system where two homologues have evolved clear and opposite functions in controlling a major developmental switch. In plants, flowering is a major developmental transition that is critical to reproductive success. Arabidopsis phosphatidylethanolamine-binding protein homologues TERMINAL FLOWER 1 (TFL1) and FLOWERING LOCUS T (FT) are key controllers of flowering, determining when and where flowers are made, but as opposing functions: TFL1 is a repressor, FT is an activator. We have uncovered a striking molecular basis for how these homologous proteins have diverged. Although <60% identical, we have shown that swapping a single amino acid is sufficient to convert TFL1 to FT function and vice versa. Therefore, these key residues may have strongly contributed to the selection of these important functions over plant evolution. Further, our results suggest that TFL1 and FT are highly conserved in biochemical function and that they act as repressors or activators of flowering through discrimination of structurally related interactors by a single residue.