Tom Viaene

Gamma paleohexaploidy in the stem-lineage of core eudicots: significance for MADS-box gene and species diversification

Comparative genome biology has unveiled the polyploid origin of all angiosperms and the role of recurrent polyploidization in the amplification of gene families and the structuring of genomes. Which species share certain ancient polyploidy events, and which do not, is ill defined because of the limited number of sequenced genomes and transcriptomes and their uneven phylogenetic distribution. Previously, it has been suggested that most, but probably not all, of the eudicots have shared an ancient hexaploidy event, referred to as the gamma triplication. In this study, detailed phylogenies of subfamilies of MADS-box genes suggest that the gamma triplication has occurred before the divergence of Gunnerales but after the divergence of Buxales and Trochodendrales. Large-scale phylogenetic and K(S)-based approaches on the inflorescence transcriptomes of Gunnera manicata (Gunnerales) and Pachysandra terminalis (Buxales) provide further support for this placement, enabling us to position the gamma triplication in the stem lineage of the core eudicots. This triplication likely initiated the functional diversification of key regulators of reproductive development in the core eudicots, comprising 75% of flowering plants. Although it is possible that the gamma event triggered early core eudicot diversification, our dating estimates suggest that the event occurred early in the stem lineage, well before the rapid speciation of the earliest core eudicot lineages. The evolutionary significance of this paleopolyploidy event may thus rather lie in establishing a species lineage that was resilient to extinction, but with the genomic potential for later diversification. We consider that the traits generated from this potential characterize extant core eudicots both chemically and morphologically.

Pistillata—Duplications as a Mode for Floral Diversification in (Basal) Asterids

Basal asterid families, and to a lesser extent the asterids as a whole, are characterized by a high variation in petal and stamen morphology. Moreover, the stamen number, the adnation of stamens to petals, and the degree of sympetaly vary considerably among basal asterid taxa. The B group genes, members of the APETALA3 (AP3) and PISTILLATA (PI) gene lineages, have been shown to specify petal and stamen identities in several core eudicot species. Duplicate genes in these lineages have been shown in some cases to have diversified in their function; for instance in Petunia, a PI paralog is required for the fusion of stamens to the corolla tube, illustrating that such genes belonging to this lineage are not just involved in specifying the identity of the stamens and petals but can also specify novel floral morphologies. This motivated us to study the duplication history of class B genes throughout asterid lineages, which comprise approximately one-third of all flowering plants. The evolutionary history of the PI gene subfamily indicates that the two genes in Petunia result from an ancient duplication event, coinciding with the origin of core asterids. A second duplication event occurred before the speciation of basal asterid Ericales families. These and other duplications in the PI lineage are not correlated with duplications in the AP3 lineage. To understand the molecular evolution of the Ericales PI genes after duplication, we have described their expression patterns using reverse transcription polymerase chain reaction and in situ hybridization, reconstructed how selection shaped their protein sequences and tested their protein interaction specificity with other class B proteins. We find that after duplication, PI paralogs have acquired multiple different expression patterns and negative selective pressure on their codons is relaxed, whereas substitutions in sites putatively involved in protein-protein interactions show positive selection, allowing for a change in the interaction behavior of the PI paralogs after duplication. Together, these observations suggest that the asterids have preferentially recruited PI duplicate genes to diverse and potentially novel roles in asterid flower development.

Expression divergence of the AGL6 MADS domain transcription factor lineage after a core eudicot duplication suggests functional diversification

BackgroundBecause of their known role as transcriptional regulators of key plant developmental processes, the diversification of MADS-box gene function is thought to be a major driving force in the developmental evolution of plants. Yet the function of some MADS-box gene subfamilies has remained elusive thus far. One such lineage, AGL6, has now been functionally characterized in three angiosperm species, but a phylogenetic framework for comparison of AGL6 gene function is currently missing.ResultsBased on phylogenetic analyses of newly isolated and EST-based sequences, we describe the duplication history of the AGL6 subfamily in angiosperms. Our analyses provide support for four ancient duplications in the evolution of the AGL6 lineage: one at the base of core eudicots resulting in euAGL6 and AGL6-like gene clades, one during basal angiosperm diversification and two in monocot evolution. To investigate whether the spatial domains in which AGL6 genes function have diverged after duplication, we use quantitative Real Time PCR. We show that the core eudicot AGL6-like clade acquired expression in vegetative tissues, while its paralog euAGL6 remains predominantly confined to reproductive tissues.ConclusionsThese and previous data lead us to propose that the AGL6 lineage in core eudicots, in addition to functions related to the expression in reproductive structures, may have acquired a function in developmental transitions of vegetative shoots.

Robustness and evolvability in the B-system of flower development.

BACKGROUNDnGene duplication has often been invoked as a key mechanism responsible for evolution of new morphologies. The floral homeotic B-group gene family has undergone a number of gene duplication events, and yet the functions of these genes appear to be largely conserved. However, detailed comparative analysis has indicated that such duplicate genes have considerable cryptic variability in their functions. In the Solanaceae, two duplicate B-group gene lineages have been retained in three subfamilies. Comparisons of orthologous genes across members of the Solanaceae have demonstrated that the combined function of all four B-gene members is to establish petal and stamen identity, but that this function was partitioned differently in each species. These observations emphasize both the robustness and the evolvability of the B-system.nnnSCOPEnWe provide an overview of how the B-function genes can robustly specify petal and stamen identity and at the same time evolve through changes in protein-protein interaction, gene expression patterns, copy number variation or alterations in the downstream genes they control. By using mathematical models we explore regulatory differences between species and how these impose constraints on downstream gene regulation.nnnCONCLUSIONSnEvolvability of the B-genes can be understood through the multiple ways in which the B-system can be robust. Quantitative approaches should allow for the incorporation of more biological realism in the representations of these regulatory systems and this should contribute to understanding the constraints under which different B-systems can function and evolve. This, in turn, can provide a better understanding of the ways in which B-genes have contributed to flower diversity.

TM8 represses developmental timing in Nicotiana benthamiana and has functionally diversified in angiosperms

BackgroundMADS-box genes are key regulators of plant reproductive development and members of most lineages of this gene family have been extensively studied. However, the function and diversification of the ancient TM8 lineage remains elusive to date. The available data suggest a possible function in flower development in tomato and fast evolution through numerous gene loss events in flowering plants.ResultsWe show the broad conservation of TM8 within angiosperms and find that in contrast to other MADS-box gene lineages, no gene duplicates have been retained after major whole genome duplication events. Through knock-down of NbTM8 by virus induced gene silencing in Nicotiana benthamiana, we show that NbTM8 represses miR172 together with another MADS-box gene, SHORT VEGETATIVE PHASE (NbSVP). In the closely related species Petunia hybrida, PhTM8 is not expressed under the conditions we investigated and consistent with this, a knock-out mutant did not show a phenotype. Finally, we generated transgenic tomato plants in which TM8 was silenced or ectopically expressed, but these plants did not display a clear phenotype. Therefore, no clear function could be confirmed for Solanum lycopersium.ConclusionsWhile the presence of TM8 is generally conserved, it remains difficult to propose a general function in angiosperms. Based on all the available data to date, supplemented with our own results, TM8 function seems to have diversified quickly throughout angiosperms and acts as repressor of miR172 in Nicotiana benthamiana, together with NbSVP.