A KNOX-Cytokinin Regulatory Module Predates the Origin of Indeterminate Vascular Plants

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Multiple innovations underpinned branching form diversification in mosses

International audienceBroad-scale evolutionary comparisons have shown that branching forms arose by convergencein vascular plants and bryophytes, but the trajectory of branching form diversificationin bryophytes is unclear. Mosses are the most species-rich bryophyte lineage andtwo sub-groups are circumscribed by alternative reproductive organ placements. In one,reproductive organs form apically, terminating growth of the primary shoot (gametophore)axis. In the other, reproductive organs develop on very short lateral branches. Aswitch from apical to lateral reproductive organ development is proposed to have primedbranching form diversification. Moss gametophores have modular development and each module develops from a singleapical cell. Here we define the architectures of 175 mosses by the number of module classes,branching patterns and the pattern in which similar modules repeat. Using ancestral characterstate reconstruction we identify two stages of architectural diversification. During a first stage there were sequential changes in the module repetition pattern, reproductiveorgan position, branching pattern and the number of module classes. During a secondstage, vegetative changes occurred independently of reproductive fate. The results pinpoint the nature of developmental change priming branching form diversificationin mosses and provide a framework for mechanistic studies of architectural diversificatio

Plasma membrane-targeted PIN proteins drive shoot development in a moss.

BACKGROUND: Plant body plans arise by the activity of meristematic growing tips during development and radiated independently in the gametophyte (n) and sporophyte (2n) stages of the life cycle during evolution. Although auxin and its intercellular transport by PIN family efflux carriers are primary regulators of sporophytic shoot development in flowering plants, the extent of conservation in PIN function within the land plants and the mechanisms regulating bryophyte gametophytic shoot development are largely unknown. RESULTS: We have found that treating gametophytic shoots of the moss Physcomitrella patens with exogenous auxins and auxin transport inhibitors disrupts apical function and leaf development. Two plasma membrane-targeted PIN proteins are expressed in leafy shoots, and pin mutants resemble plants treated with auxins or auxin transport inhibitors. PIN-mediated auxin transport regulates apical cell function, leaf initiation, leaf shape, and shoot tropisms in moss gametophytes. pin mutant sporophytes are sometimes branched, reproducing a phenotype only previously seen in the fossil record and in rare natural moss variants. CONCLUSIONS: Our results show that PIN-mediated auxin transport is an ancient, conserved regulator of shoot development.C.J.H. is funded by a Royal Society University Research Fellowship, a Gatsby Charitable Foundation fellowship (GAT2962) and the BBSRC (BB/L00224811) and R.R. is funded by the Deutsche Forschungsgemeinschaft (SPP 1067, RE 837/6) and the Excellence Initiative of the German Federal and State Governments (EXC294).This is the final version. It was first published by Elsevier at http://www.cell.com/current-biology/abstract/S0960-9822%2814%2901217-2

Three non-autonomous signals collaborate for nuclear targeting of CrMYC2, a Catharanthus roseus bHLH transcription factor

<p>Abstract</p> <p>Background</p> <p>CrMYC2 is an early jasmonate-responsive bHLH transcription factor involved in the regulation of the expression of the genes of the terpenic indole alkaloid biosynthesis pathway in <it>Catharanthus roseus</it>. In this paper, we identified the amino acid domains necessary for the nuclear targeting of CrMYC2.</p> <p>Findings</p> <p>We examined the intracellular localization of whole CrMYC2 and of various deletion mutants, all fused with GFP, using a transient expression assay in onion epidermal cells. Sequence analysis of this protein revealed the presence of four putative basic nuclear localization signals (NLS). Assays showed that none of the predicted NLS is active alone. Further functional dissection of CrMYC2 showed that the nuclear targeting of this transcription factor involves the cooperation of three domains located in the C-terminal region of the protein. The first two domains are located at amino acid residues 454-510 and 510-562 and contain basic classical monopartite NLSs; these regions are referred to as NLS3 (KRPRKR) and NLS4 (EAERQRREK), respectively. The third domain, between residues 617 and 652, is rich in basic amino acids that are well conserved in other phylogenetically related bHLH transcription factors. Our data revealed that these three domains are inactive when isolated but act cooperatively to target CrMYC2 to the nucleus.</p> <p>Conclusions</p> <p>This study identified three amino acid domains that act in cooperation to target the CrMYC2 transcription factor to the nucleus. Further fine structure/function analysis of these amino acid domains will allow the identification of new NLS domains and will allow the investigation of the related molecular mechanisms involved in the nuclear targeting of the CrMYC2 bHLH transcription factor.</p

Functional study of CROWN ROOTLESS1, a AS2/LOB domain protein essential for rice crown root development

Chez le riz, la céréale modèle, le système racinaire est principalement constitué de racines issues de la tige, nommées racines coronaires (RC). Peu de gènes contrôlant le développement des RC sont connus, parmi eux CROWN ROOTLESS1 (CRL1) code une protéine à domaine AS2/LOB (ASL/LBD), qui est probablement un facteur de transcription. Le gène CRL1 est nécessaire à l'initiation des primordia de RC, il est directement activé par l'auxine et est situé en amont du réseau de gènes contrôlant le programme de différentiation des RC. Afin de mieux connaître les processus génétiques impliqués dans l'initiation des RC, l!objectif principal de cette thèse est de comprendre la fonction moléculaire de la protéine CRL1 en validant sa fonction de facteur de transcription et en identifiant ses gènes cibles. L'interaction de la protéine CRL1 avec l!ADN a été montrée in vitro et une expérience de SELEX a permis d'identifier sa séquence de fixation à l!ADN : CACA(A/C)C (CRL1-box). Des expériences en levure ont permis de montrer que CRL1 est un activateur de la transcription. Une comparaison entre le sauvage et le mutant crl1, ainsi que l'élaboration d!un système inductible à la dexaméthasone permettant d'activer l'expression de CRL1 dans le fond génétique mutant crl1, ont été utilisés pour identifier des gènes cibles précoces de CRL1 grâce à des analyse de transcriptome. 277 gènes sont activés dès quatre heures après induction de CRL1, les deux tiers contiennent au moins une CRL1-box dans leur promoteur et peuvent donc être des cibles directes de CRL1. CRL1 induit l'expression d!un ensemble de gènes permettant la mise en place des processus de régulation de l'information génétique, de division, de croissance et de différenciation cellulaires nécessaires à la création d'un méristème de racine coronaire organisé et fonctionnel. Parmi eux, QHB code un facteur de transcription clé nécessaire au maintien des cellules souches des méristèmes racinaires. Ce résultat établit pour la première fois un lien moléculaire entre la signalisation de l!auxine et des gènes impliqués dans la mise en place ou le maintien des cellules souches lors de la formation d!un nouveau méristème racinaire au cours du développement post-embryonnaire. Par ailleurs, une étude histologique a permis de révéler que les RC sont issues d!une couche de péricycle dans la tige, un tissu équivalent en termes de localisation et de potentiel rhizogène au péricycle de la racine à partir duquel sont initiées les racines latérales. Les données acquises suggèrent de fortes similarités dans les processus cellulaires et génétiques de la différentiation des méristèmes racinaires au cours du développement post-embryonnaire chez les monocotylédones et les dicotylédones. La découverte de gènes spécifiques au développement de racines issues de la tige ouvre une voie importante vers la compréhension du déterminisme génétique de l!architecture du système racinaire chez les céréales et offre un nouveau potentiel de ressources génétiques pour l!amélioration variétale.In rice, the model cereal, the root system is mainly composed of stem-derived roots, named crown roots (CR). Very few genes that control the root system development are known, among them CROWN ROOTLESS1 (CRL1) encodes an AS2/LOB-domain protein that is a putative transcription factor (TF). CRL1 is necessary for CR primordium initiation, it is directly activated by auxin and is situated upstream of the gene regulatory network that control the CR differentiation programme. To better known the genetic processes involved in CR initiation, the main objective of this thesis is to understand the molecular function of the CRL1 protein by validating its function of TF and by identifying its target genes. The interaction of CRL1 with DNA was shown in vitro and a consensus CRL1 DNA-binding motif was identified with a SELEX method : CACA(A/C)C named CRL1-box. A yeast assay showed that CRL1 is a transcriptional activator. A comparison between wild type and c rl1 mutant, and the development of a dexamethasone inducible system to ectopically express CRL1 in the crl1 background, were used to identify CRL1 early target genes by transcript profiling. 277 genes were induced from four hours following CRL1 activation, the two-thirds possess at least one CRL1-box and may be CRL1 direct target genes. CRL1 activates the expression of a broad range of genes that allow to orientate genome expression and to initiate cell division, growth and differentiation mechanisms required for the building of an organized and functional CR meristem. Among these genes, QHB encodes a key TF required for the maintenance of root meristem stem cells. This result evidences for the first time a molecular link between auxin signalling and major genes involved in stem cell patterning and maintenance in the formation of a new root meristem during post-embryonic development. Otherwise, an histological study showed that CR are derived from a shoot pericycle, a tissue equivalent to the root pericycle, from which lateral roots develop, in terms of location and rhizogenic potential. All these data suggest strong similarities between monocots and dicots in cellular and genetic mechanisms that control root meristem differentiation during post-embryonic development. The identification of genes specifically involved in stem-derived root development pave the way towards the understanding of the genetic control of root system architecture in cereals and offer a new potential of genetic resources for plant breeding

The Evolution of Branching in Land Plants: Between Conservation and Diversity

International audienceThe evolution of branching was pivotal to the diversification of plant architecture, providing ways to colonize the environment and optimize resource acquisition both above and below ground. Fossil and phylogenetic evidence indicates that branching evolved independently in the two generations of the land plant life cycle. In this chapter, I focus on shoot systems and discuss two contrasting patterns, occurring at different levels: conservation and diversity. I show that two similar branching modes, terminal and lateral, are found across extant and extinct land plant lineages. Despite conservation of gross morphology, branching morphogenesis is highly disparate at the cellular level and is orchestrated in various manners between and within lineages. In contrast, the molecular mechanisms underpinning branch development could be largely shared among land plants

Etude fonctionnelle de CROWNROOTLESS1, une protéine à domaine AS2/LOB nécessaire au développement des racines coronnaires chez le riz

Chez le riz, la céréale modèle, le système racinaire est principalement constitué de racines issues de la tige, nommées racines coronaires (RC). Peu de gènes contrôlant le développement des RC sont connus, parmi eux CROWN ROOTLESS1 (CRL1) code une protéine à domaine AS2/LOB (ASL/LBD), qui est probablement un facteur de transcription. Le gène CRL1 est nécessaire à l'initiation des primordia de RC, il est directement activé par l'auxine et est situé en amont du réseau de gènes contrôlant le programme de différentiation des RC. Afin de mieux connaître les processus génétiques impliqués dans l'initiation des RC, l!objectif principal de cette thèse est de comprendre la fonction moléculaire de la protéine CRL1 en validant sa fonction de facteur de transcription et en identifiant ses gènes cibles. L'interaction de la protéine CRL1 avec l!ADN a été montrée in vitro et une expérience de SELEX a permis d'identifier sa séquence de fixation à l!ADN : CACA(A/C)C (CRL1-box). Des expériences en levure ont permis de montrer que CRL1 est un activateur de la transcription. Une comparaison entre le sauvage et le mutant crl1, ainsi que l'élaboration d!un système inductible à la dexaméthasone permettant d'activer l'expression de CRL1 dans le fond génétique mutant crl1, ont été utilisés pour identifier des gènes cibles précoces de CRL1 grâce à des analyse de transcriptome. 277 gènes sont activés dès quatre heures après induction de CRL1, les deux tiers contiennent au moins une CRL1-box dans leur promoteur et peuvent donc être des cibles directes de CRL1. CRL1 induit l'expression d!un ensemble de gènes permettant la mise en place des processus de régulation de l'information génétique, de division, de croissance et de différenciation cellulaires nécessaires à la création d'un méristème de racine coronaire organisé et fonctionnel. Parmi eux, QHB code un facteur de transcription clé nécessaire au maintien des cellules souches des méristèmes racinaires. Ce résultat établit pour la première fois un lien moléculaire entre la signalisation de l!auxine et des gènes impliqués dans la mise en place ou le maintien des cellules souches lors de la formation d!un nouveau méristème racinaire au cours du développement post-embryonnaire. Par ailleurs, une étude histologique a permis de révéler que les RC sont issues d!une couche de péricycle dans la tige, un tissu équivalent en termes de localisation et de potentiel rhizogène au péricycle de la racine à partir duquel sont initiées les racines latérales. Les données acquises suggèrent de fortes similarités dans les processus cellulaires et génétiques de la différentiation des méristèmes racinaires au cours du développement post-embryonnaire chez les monocotylédones et les dicotylédones. La découverte de gènes spécifiques au développement de racines issues de la tige ouvre une voie importante vers la compréhension du déterminisme génétique de l!architecture du système racinaire chez les céréales et offre un nouveau potentiel de ressources génétiques pour l!amélioration variétale.In rice, the model cereal, the root system is mainly composed of stem-derived roots, named crown roots (CR). Very few genes that control the root system development are known, among them CROWN ROOTLESS1 (CRL1) encodes an AS2/LOB-domain protein that is a putative transcription factor (TF). CRL1 is necessary for CR primordium initiation, it is directly activated by auxin and is situated upstream of the gene regulatory network that control the CR differentiation programme. To better known the genetic processes involved in CR initiation, the main objective of this thesis is to understand the molecular function of the CRL1 protein by validating its function of TF and by identifying its target genes. The interaction of CRL1 with DNA was shown in vitro and a consensus CRL1 DNA-binding motif was identified with a SELEX method : CACA(A/C)C named CRL1-box. A yeast assay showed that CRL1 is a transcriptional activator. A comparison between wild type and c rl1 mutant, and the development of a dexamethasone inducible system to ectopically express CRL1 in the crl1 background, were used to identify CRL1 early target genes by transcript profiling. 277 genes were induced from four hours following CRL1 activation, the two-thirds possess at least one CRL1-box and may be CRL1 direct target genes. CRL1 activates the expression of a broad range of genes that allow to orientate genome expression and to initiate cell division, growth and differentiation mechanisms required for the building of an organized and functional CR meristem. Among these genes, QHB encodes a key TF required for the maintenance of root meristem stem cells. This result evidences for the first time a molecular link between auxin signalling and major genes involved in stem cell patterning and maintenance in the formation of a new root meristem during post-embryonic development. Otherwise, an histological study showed that CR are derived from a shoot pericycle, a tissue equivalent to the root pericycle, from which lateral roots develop, in terms of location and rhizogenic potential. All these data suggest strong similarities between monocots and dicots in cellular and genetic mechanisms that control root meristem differentiation during post-embryonic development. The identification of genes specifically involved in stem-derived root development pave the way towards the understanding of the genetic control of root system architecture in cereals and offer a new potential of genetic resources for plant breeding.MONTPELLIER-BU Sciences (341722106) / SudocSudocFranceF

Re-examining meristems through the lens of evo-devo

International audienceThe concept of the meristem was introduced in 1858 to characterize multicellular, formative, and proliferative tissues that give rise to the entire plant body, based on observations of vascular plants. Although its original definition did not encompass bryophytes, this concept has been used and continuously refined over the past 165 years to describe the diverse apices of all land plants. Here, we re-examine this matter in light of recent evo-devo research and show that, despite displaying high anatomical diversity, land plant meristems are unified by shared genetic control. We also propose a modular view of meristem function and highlight multiple evolutionary mechanisms that are likely to have contributed to the assembly and diversification of the varied meristems during the course of plant evolution

Phyllotaxis from a Single Apical Cell

International audiencePhyllotaxis, the geometry of leaf arrangement around stems, determines plant architecture. Molecular interactions coordinating the formation of phyllotactic patterns have mainly been studied in multicellular shoot apical meristems of flowering plants. Phyllotaxis evolved independently in the major land plant lineages. In mosses, it arises from a single apical cell, raising the question of how asymmetric divisions of a single-celled meristem create phyllotactic patterns and whether associated genetic processes are shared across lineages. We present an overview of the mechanisms governing shoot apical cell specification and activity in the model moss, Physcomitrium patens, and argue that similar molecular regulatory modules have been deployed repeatedly across evolution to operate at different scales and driv