Publikationen - Molekulare Signalverarbeitung

Combining transcriptome data of biological processes or response to stimuli with evolutionary information such as the phylogenetic conservation of genes or their sequence divergence rates enables the investigation of evolutionary constraints on these processes or responses. Such phylotranscriptomic analyses recently unraveled that mid-developmental transcriptomes of fly, fish, and cress were dominated by evolutionarily conserved genes and genes under negative selection and thus recapitulated the developmental hourglass on the transcriptomic level. Here, we present a protocol for performing phylotranscriptomic analyses on any biological process of interest. When applying this protocol, users are capable of detecting different evolutionary constraints acting on different stages of the biological process of interest in any species. For each step of the protocol, modular and easy-to-use open-source software tools are provided, which enable a broad range of scientists to apply phylotranscriptomic analyses to a wide spectrum of biological questions.

The historic developmental hourglass concept depicts the convergence of animal embryos to a common form during the phylotypic period. Recently, it has been shown that a transcriptomic hourglass is associated with this morphological pattern, consistent with the idea of underlying selective constraints due to intense molecular interactions during body plan establishment. Although plants do not exhibit a morphological hourglass during embryogenesis, a transcriptomic hourglass has nevertheless been identified in the model plant Arabidopsis thaliana. Here, we investigated whether plant hourglass patterns are also found postembryonically. We found that the two main phase changes during the life cycle of Arabidopsis, from embryonic to vegetative and from vegetative to reproductive development, are associated with transcriptomic hourglass patterns. In contrast, flower development, a process dominated by organ formation, is not. This suggests that plant hourglass patterns are decoupled from organogenesis and body plan establishment. Instead, they may reflect general transitions through organizational checkpoints.

The historic developmental hourglass concept depicts the convergence of animal embryos to a common form during the phylotypic period. Recently, it has been shown that a transcriptomic hourglass is associated with this morphological pattern, consistent with the idea of underlying selective constraints due to intense molecular interactions during body plan establishment. Although plants do not exhibit a morphological hourglass during embryogenesis, a transcriptomic hourglass has nevertheless been identified in the model plant Arabidopsis thaliana. Here, we investigated whether plant hourglass patterns are also found post-embryonically. We found that the two main phase changes during the life cycle of Arabidopsis, from embryonic to vegetative and from vegetative to reproductive development, are associated with transcriptomic hourglass patterns. In contrast, flower development, a process dominated by organ formation, is not. This suggests that plant hourglass patterns are decoupled from organogenesis and body plan establishment. Instead, they may reflect general transitions through organizational checkpoints.

The developmental hourglass model has been used to describe the morphological transitions of related species throughout embryogenesis. Recently, quantifiable approaches combining transcriptomic and evolutionary information provided novel evidence for the presence of a phylotranscriptomic hourglass pattern across kingdoms. As its biological function is unknown it remains speculative whether this pattern is functional or merely represents a nonfunctional evolutionary relic. The latter would seriously hamper future experimental approaches designed to test hypotheses regarding its function. Here, we address this question by generating transcriptome divergence index (TDI) profiles across embryogenesis of Danio rerio, Drosophila melanogaster, and Arabidopsis thaliana. To enable meaningful evaluation of the resulting patterns, we develop a statistical test that specifically assesses potential hourglass patterns. Based on this objective measure we find that two of these profiles follow a statistically significant hourglass pattern with the most conserved transcriptomes in the phylotypic periods. As the TDI considers only recent evolutionary signals, this indicates that the phylotranscriptomic hourglass pattern is not a rudiment but possibly actively maintained, implicating the existence of some linked biological function associated with embryogenesis in extant species.

BackgroundThe formation of flowers is one of the main model systems to elucidate the molecular mechanisms that control developmental processes in plants. Although several studies have explored gene expression during flower development in the model plant Arabidopsis thaliana on a genome-wide scale, a continuous series of expression data from the earliest floral stages until maturation has been lacking. Here, we used a floral induction system to close this information gap and to generate a reference dataset for stage-specific gene expression during flower formation.ResultsUsing a floral induction system, we collected floral buds at 14 different stages from the time of initiation until maturation. Using whole-genome microarray analysis, we identified 7,405 genes that exhibit rapid expression changes during flower development. These genes comprise many known floral regulators and we found that the expression profiles for these regulators match their known expression patterns, thus validating the dataset. We analyzed groups of co-expressed genes for over-represented cellular and developmental functions through Gene Ontology analysis and found that they could be assigned specific patterns of activities, which are in agreement with the progression of flower development. Furthermore, by mapping binding sites of floral organ identity factors onto our dataset, we were able to identify gene groups that are likely predominantly under control of these transcriptional regulators. We further found that the distribution of paralogs among groups of co-expressed genes varies considerably, with genes expressed predominantly at early and intermediate stages of flower development showing the highest proportion of such genes.ConclusionsOur results highlight and describe the dynamic expression changes undergone by a large number of genes during flower development. They further provide a comprehensive reference dataset for temporal gene expression during flower formation and we demonstrate that it can be used to integrate data from other genomics approaches such as genome-wide localization studies of transcription factor binding sites.

The plant hormone auxin regulates virtually every aspect of plant growth and development. Auxin acts by binding the F-box protein transport inhibitor response 1 (TIR1) and promotes the degradation of the AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) transcriptional repressors. Here we show that efficient auxin binding requires assembly of an auxin co-receptor complex consisting of TIR1 and an Aux/IAA protein. Heterologous experiments in yeast and quantitative IAA binding assays using purified proteins showed that different combinations of TIR1 and Aux/IAA proteins form co-receptor complexes with a wide range of auxin-binding affinities. Auxin affinity seems to be largely determined by the Aux/IAA. As there are 6 TIR1/AUXIN SIGNALING F-BOX proteins (AFBs) and 29 Aux/IAA proteins in Arabidopsis thaliana, combinatorial interactions may result in many co-receptors with distinct auxin-sensing properties. We also demonstrate that the AFB5–Aux/IAA co-receptor selectively binds the auxinic herbicide picloram. This co-receptor system broadens the effective concentration range of the hormone and may contribute to the complexity of auxin response.

Animal and plant development starts with a constituting phase called embryogenesis, which evolved independently in both lineages1. Comparative anatomy of vertebrate development—based on the Meckel-Serrès law2 and von Baer’s laws of embryology3 from the early nineteenth century—shows that embryos from various taxa appear different in early stages, converge to a similar form during mid-embryogenesis, and again diverge in later stages. This morphogenetic series is known as the embryonic ‘hourglass’4,5, and its bottleneck of high conservation in mid-embryogenesis is referred to as the phylotypic stage6. Recent analyses in zebrafish and Drosophila embryos provided convincing molecular support for the hourglass model, because during the phylotypic stage the transcriptome was dominated by ancient genes7 and global gene expression profiles were reported to be most conserved8. Although extensively explored in animals, an embryonic hourglass has not been reported in plants, which represent the second major kingdom in the tree of life that evolved embryogenesis. Here we provide phylotranscriptomic evidence for a molecular embryonic hourglass in Arabidopsis thaliana, using two complementary approaches. This is particularly significant because the possible absence of an hourglass based on morphological features in plants suggests that morphological and molecular patterns might be uncoupled. Together with the reported developmental hourglass patterns in animals, these findings indicate convergent evolution of the molecular hourglass and a conserved logic of embryogenesis across kingdoms.

The identity of the auxin receptor(s) and the mechanism of auxin perception has been a subject of intense interest since the discovery of auxin almost a century ago. The development of genetic approaches to the study of plant hormone signaling led to the discovery that auxin acts by promoting degradation of transcriptional repressors called Aux/IAA proteins. This process requires a ubiquitin protein ligase (E3) called SCFTIR1 and related SCF complexes. Surprisingly, auxin works by directly binding to TIR1, the F-box protein subunit of this SCF. Structural studies demonstrate that auxin acts like a “molecular glue,” to stabilize the interaction between TIR1 and the Aux/IAA substrate. These exciting results solve an old problem in plant biology and reveal new mechanisms for E3 regulation and hormone perception.

Auxin is a pivotal plant hormone that controls many aspects of plant growth and development. Perceived by a small family of F-box proteins including transport inhibitor response 1 (TIR1), auxin regulates gene expression by promoting SCF ubiquitin-ligase-catalysed degradation of the Aux/IAA transcription repressors, but how the TIR1 F-box protein senses and becomes activated by auxin remains unclear. Here we present the crystal structures of the Arabidopsis TIR1–ASK1 complex, free and in complexes with three different auxin compounds and an Aux/IAA substrate peptide. These structures show that the leucine-rich repeat domain of TIR1 contains an unexpected inositol hexakisphosphate co-factor and recognizes auxin and the Aux/IAA polypeptide substrate through a single surface pocket. Anchored to the base of the TIR1 pocket, auxin binds to a partially promiscuous site, which can also accommodate various auxin analogues. Docked on top of auxin, the Aux/IAA substrate peptide occupies the rest of the TIR1 pocket and completely encloses the hormone-binding site. By filling in a hydrophobic cavity at the protein interface, auxin enhances the TIR1–substrate interactions by acting as a ‘molecular glue’. Our results establish the first structural model of a plant hormone receptor.