Publications by authors named "François Parcy"

62 Publications

The LEAFY floral regulator displays pioneer transcription factor properties.

Mol Plant 2021 Mar 5. Epub 2021 Mar 5.

Laboratoire Physiologie Cellulaire et Végétale, Université Grenoble Alpes, CNRS, CEA, INRAE, IRIG-DBSCI-LPCV, 17 avenue des martyrs, 38054, Grenoble, France. Electronic address:

Pioneer transcription factors (TFs) are a special category of TFs with the capacity to bind to closed chromatin regions in which DNA is wrapped around histones and may be highly methylated. Subsequently, pioneer TFs are able to modify the chromatin state to initiate gene expression. In plants, LEAFY (LFY) is a master floral regulator and has been suggested to act as a pioneer TF in Arabidopsis. Here, we demonstrate that LFY is able to bind both methylated and non-methylated DNA using a combination of in vitro genome-wide binding experiments and structural modeling. Comparisons between regions bound by LFY in vivo and chromatin accessibility data suggest that a subset of LFY bound regions is occupied by nucleosomes. We confirm that LFY is able to bind nucleosomal DNA in vitro using reconstituted nucleosomes. Finally, we show that constitutive LFY expression in seedling tissues is sufficient to induce chromatin accessibility in the LFY direct target genes APETALA1 and AGAMOUS. Taken together, our study suggests that LFY possesses key pioneer TF features that contribute to launching the floral gene expression program.
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http://dx.doi.org/10.1016/j.molp.2021.03.004DOI Listing
March 2021

A network of transcriptional repressors modulates auxin responses.

Nature 2021 01 18;589(7840):116-119. Epub 2020 Nov 18.

Laboratoire Reproduction et Développement des Plantes, Université de Lyon, ENS de Lyon, UCB Lyon 1, CNRS, INRAE, Lyon, France.

The regulation of signalling capacity, combined with the spatiotemporal distribution of developmental signals themselves, is pivotal in setting developmental responses in both plants and animals. The hormone auxin is a key signal for plant growth and development that acts through the AUXIN RESPONSE FACTOR (ARF) transcription factors. A subset of these, the conserved class A ARFs, are transcriptional activators of auxin-responsive target genes that are essential for regulating auxin signalling throughout the plant lifecycle. Although class A ARFs have tissue-specific expression patterns, how their expression is regulated is unknown. Here we show, by investigating chromatin modifications and accessibility, that loci encoding these proteins are constitutively open for transcription. Through yeast one-hybrid screening, we identify the transcriptional regulators of the genes encoding class A ARFs from Arabidopsis thaliana and demonstrate that each gene is controlled by specific sets of transcriptional regulators. Transient transformation assays and expression analyses in mutants reveal that, in planta, the majority of these regulators repress the transcription of genes encoding class A ARFs. These observations support a scenario in which the default configuration of open chromatin enables a network of transcriptional repressors to regulate expression levels of class A ARF proteins and modulate auxin signalling output throughout development.
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http://dx.doi.org/10.1038/s41586-020-2940-2DOI Listing
January 2021

Genome-wide binding of SEPALLATA3 and AGAMOUS complexes determined by sequential DNA-affinity purification sequencing.

Nucleic Acids Res 2020 09;48(17):9637-9648

Laboratoire de Physiologie Cellulaire et Végétale, Université Grenoble-Alpes, CNRS, CEA, INRAE, IRIG-DBSCI, 38000 Grenoble, France.

The MADS transcription factors (TF), SEPALLATA3 (SEP3) and AGAMOUS (AG) are required for floral organ identity and floral meristem determinacy. While dimerization is obligatory for DNA binding, SEP3 and SEP3-AG also form tetrameric complexes. How homo and hetero-dimerization and tetramerization of MADS TFs affect genome-wide DNA-binding and gene regulation is not known. Using sequential DNA affinity purification sequencing (seq-DAP-seq), we determined genome-wide binding of SEP3 homomeric and SEP3-AG heteromeric complexes, including SEP3Δtet-AG, a complex with a SEP3 splice variant, SEP3Δtet, which is largely dimeric and SEP3-AG tetramer. SEP3 and SEP3-AG share numerous bound regions, however each complex bound unique sites, demonstrating that protein identity plays a role in DNA-binding. SEP3-AG and SEP3Δtet-AG share a similar genome-wide binding pattern; however the tetrameric form could access new sites and demonstrated a global increase in DNA-binding affinity. Tetramerization exhibited significant cooperative binding with preferential distances between two sites, allowing efficient binding to regions that are poorly recognized by dimeric SEP3Δtet-AG. By intersecting seq-DAP-seq with ChIP-seq and expression data, we identified unique target genes bound either in SEP3-AG seq-DAP-seq or in SEP3/AG ChIP-seq. Seq-DAP-seq is a versatile genome-wide technique and complements in vivo methods to identify putative direct regulatory targets.
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http://dx.doi.org/10.1093/nar/gkaa729DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7515736PMC
September 2020

OsFD4 promotes the rice floral transition via florigen activation complex formation in the shoot apical meristem.

New Phytol 2021 01 27;229(1):429-443. Epub 2020 Aug 27.

Department of Biosciences, University of Milan, Milan, 20123, Italy.

In rice, the florigens Heading Date 3a (Hd3a) and Rice Flowering Locus T 1 (RFT1), OsFD-like basic leucine zipper (bZIP) transcription factors, and Gf14 proteins assemble into florigen activation/repressor complexes (FACs/FRCs), which regulate transition to flowering in leaves and apical meristem. Only OsFD1 has been described as part of complexes promoting flowering at the meristem, and little is known about the role of other bZIP transcription factors, the combinatorial complexity of FAC formation, and their DNA-binding properties. Here, we used mutant analysis, protein-protein interaction assays and DNA affinity purification (DAP) sequencing coupled to in silico prediction of binding syntaxes to study several bZIP proteins that assemble into FACs or FRCs. We identified OsFD4 as a component of a FAC promoting flowering at the shoot apical meristem, downstream of OsFD1. The osfd4 mutants are late flowering and delay expression of genes promoting inflorescence development. Protein-protein interactions indicate an extensive network of contacts between several bZIPs and Gf14 proteins. Finally, we identified genomic regions bound by bZIPs with promotive and repressive effects on flowering. We conclude that distinct bZIPs orchestrate floral induction at the meristem and that FAC formation is largely combinatorial. While binding to the same consensus motif, their DNA-binding syntax is different, suggesting discriminatory functions.
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http://dx.doi.org/10.1111/nph.16834DOI Listing
January 2021

Contrasted evolutionary trajectories of plant transcription factors.

Curr Opin Plant Biol 2020 04 15;54:101-107. Epub 2020 May 15.

Laboratoire Physiologie Cellulaire et Végétale, Université Grenoble Alpes, CNRS, CEA, INRAE, IRIG-LPCV, 38000 Grenoble, France. Electronic address:

Because of their prominent roles in plant development, transcription factors (TF) play central roles as drivers of innovation in the evolution of the green lineage (viridiplantae). The advent of massive sequencing combined with comparative genetics/genomics allows a rigorous investigation of how TF families have contributed to plant diversification from charophyte algae to bryophytes to angiosperms. Here, we review recent progress on TF family reconstruction and the identification of distantly related TFs present throughout the evolutionary timeline from algae to angiosperms. These data provide examples of contrasting evolutionary trajectories of TF families and illustrate how conserved TFs adopt diverse roles over the course of evolution.
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http://dx.doi.org/10.1016/j.pbi.2020.03.002DOI Listing
April 2020

JASPAR 2020: update of the open-access database of transcription factor binding profiles.

Nucleic Acids Res 2020 01;48(D1):D87-D92

Centre for Molecular Medicine Norway (NCMM), Nordic EMBL Partnership, University of Oslo, 0318 Oslo, Norway.

JASPAR (http://jaspar.genereg.net) is an open-access database of curated, non-redundant transcription factor (TF)-binding profiles stored as position frequency matrices (PFMs) for TFs across multiple species in six taxonomic groups. In this 8th release of JASPAR, the CORE collection has been expanded with 245 new PFMs (169 for vertebrates, 42 for plants, 17 for nematodes, 10 for insects, and 7 for fungi), and 156 PFMs were updated (125 for vertebrates, 28 for plants and 3 for insects). These new profiles represent an 18% expansion compared to the previous release. JASPAR 2020 comes with a novel collection of unvalidated TF-binding profiles for which our curators did not find orthogonal supporting evidence in the literature. This collection has a dedicated web form to engage the community in the curation of unvalidated TF-binding profiles. Moreover, we created a Q&A forum to ease the communication between the user community and JASPAR curators. Finally, we updated the genomic tracks, inference tool, and TF-binding profile similarity clusters. All the data is available through the JASPAR website, its associated RESTful API, and through the JASPAR2020 R/Bioconductor package.
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http://dx.doi.org/10.1093/nar/gkz1001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7145627PMC
January 2020

Crystal structure of the transcriptional repressor DdrO: insight into the metalloprotease/repressor-controlled radiation response in Deinococcus.

Nucleic Acids Res 2019 12;47(21):11403-11417

Aix Marseille Univ, CEA, CNRS, BIAM, Molecular and Environmental Microbiology Team, Saint Paul-Lez-Durance, F-13108, France.

Exposure to harmful conditions such as radiation and desiccation induce oxidative stress and DNA damage. In radiation-resistant Deinococcus bacteria, the radiation/desiccation response is controlled by two proteins: the XRE family transcriptional repressor DdrO and the COG2856 metalloprotease IrrE. The latter cleaves and inactivates DdrO. Here, we report the biochemical characterization and crystal structure of DdrO, which is the first structure of a XRE protein targeted by a COG2856 protein. DdrO is composed of two domains that fold independently and are separated by a flexible linker. The N-terminal domain corresponds to the DNA-binding domain. The C-terminal domain, containing three alpha helices arranged in a novel fold, is required for DdrO dimerization. Cleavage by IrrE occurs in the loop between the last two helices of DdrO and abolishes dimerization and DNA binding. The cleavage site is hidden in the DdrO dimer structure, indicating that IrrE cleaves DdrO monomers or that the interaction with IrrE induces a structural change rendering accessible the cleavage site. Predicted COG2856/XRE regulatory protein pairs are found in many bacteria, and available data suggest two different molecular mechanisms for stress-induced gene expression: COG2856 protein-mediated cleavage or inhibition of oligomerization without cleavage of the XRE repressor.
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http://dx.doi.org/10.1093/nar/gkz883DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6868357PMC
December 2019

Evolution of the Auxin Response Factors from charophyte ancestors.

PLoS Genet 2019 09 25;15(9):e1008400. Epub 2019 Sep 25.

Univ. Grenoble Alpes, CNRS, CEA, INRA, IRIG-DBSCI-LPCV, Grenoble, France.

Auxin is a major developmental regulator in plants and the acquisition of a transcriptional response to auxin likely contributed to developmental innovations at the time of water-to-land transition. Auxin Response Factors (ARFs) Transcription Factors (TFs) that mediate auxin-dependent transcriptional changes are divided into A, B and C evolutive classes in land plants. The origin and nature of the first ARF proteins in algae is still debated. Here, we identify the most 'ancient' ARF homologue to date in the early divergent charophyte algae Chlorokybus atmophyticus, CaARF. Structural modelling combined with biochemical studies showed that CaARF already shares many features with modern ARFs: it is capable of oligomerization, interacts with the TOPLESS co-repressor and specifically binds Auxin Response Elements as dimer. In addition, CaARF possesses a DNA-binding specificity that differs from class A and B ARFs and that was maintained in class C ARF along plants evolution. Phylogenetic evidence together with CaARF biochemical properties indicate that the different classes of ARFs likely arose from an ancestral proto-ARF protein with class C-like features. The foundation of auxin signalling would have thus happened from a pre-existing hormone-independent transcriptional regulation together with the emergence of a functional hormone perception complex.
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http://dx.doi.org/10.1371/journal.pgen.1008400DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6797205PMC
September 2019

Building Transcription Factor Binding Site Models to Understand Gene Regulation in Plants.

Mol Plant 2019 06 15;12(6):743-763. Epub 2018 Nov 15.

CNRS, Univ. Grenoble Alpes, CEA, INRA, BIG-LPCV, 38000 Grenoble, France. Electronic address:

Transcription factors (TFs) are key cellular components that control gene expression. They recognize specific DNA sequences, the TF binding sites (TFBSs), and thus are targeted to specific regions of the genome where they can recruit transcriptional co-factors and/or chromatin regulators to fine-tune spatiotemporal gene regulation. Therefore, the identification of TFBSs in genomic sequences and their subsequent quantitative modeling is of crucial importance for understanding and predicting gene expression. Here, we review how TFBSs can be determined experimentally, how the TFBS models can be constructed in silico, and how they can be optimized by taking into account features such as position interdependence within TFBSs, DNA shape, and/or by introducing state-of-the-art computational algorithms such as deep learning methods. In addition, we discuss the integration of context variables into the TFBS modeling, including nucleosome positioning, chromatin states, methylation patterns, 3D genome architectures, and TF cooperative binding, in order to better predict TF binding under cellular contexts. Finally, we explore the possibilities of combining the optimized TFBS model with technological advances, such as targeted TFBS perturbation by CRISPR, to better understand gene regulation, evolution, and plant diversity.
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http://dx.doi.org/10.1016/j.molp.2018.10.010DOI Listing
June 2019

Capturing Auxin Response Factors Syntax Using DNA Binding Models.

Mol Plant 2019 06 15;12(6):822-832. Epub 2018 Oct 15.

Univ. Grenoble Alpes, CNRS, CEA, INRA, BIG-LPCV, 38000 Grenoble, France. Electronic address:

Auxin is a key hormone performing a wealth of functions throughout the life cycle of plants. It acts largely by regulating genes at the transcriptional level through a family of transcription factors called auxin response factors (ARFs). Even though all ARF monomers analyzed so far bind a similar DNA sequence, there is evidence that ARFs differ in their target genomic regions and regulated genes. Here, we report the use of position weight matrices (PWMs) to model ARF DNA binding specificity based on published DNA affinity purification sequencing (DAP-seq) data. We found that the genome binding of two ARFs (ARF2 and ARF5/Monopteros [MP]) differ largely because these two factors have different preferred ARF binding site (ARFbs) arrangements (orientation and spacing). We illustrated why PWMs are more versatile to reliably identify ARFbs than the widely used consensus sequences and demonstrated their power with biochemical experiments in the identification of the regulatory regions of IAA19, an well-characterized auxin-responsive gene. Finally, we combined gene regulation by auxin with ARF-bound regions and identified specific ARFbs configurations that are over-represented in auxin-upregulated genes, thus deciphering the ARFbs syntax functional for regulation. Our study provides a general method to exploit the potential of genome-wide DNA binding assays and to decode gene regulation.
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http://dx.doi.org/10.1016/j.molp.2018.09.010DOI Listing
June 2019

LEAFY activity is post-transcriptionally regulated by BLADE ON PETIOLE2 and CULLIN3 in Arabidopsis.

New Phytol 2018 10 11;220(2):579-592. Epub 2018 Jul 11.

Univ. Grenoble Alpes, CNRS, CEA, INRA, BIG-LPCV, 38054, Grenoble Cedex 9, France.

The Arabidopsis LEAFY (LFY) transcription factor is a key regulator of floral meristem emergence and identity. LFY interacts genetically and physically with UNUSUAL FLORAL ORGANS, a substrate adaptor of CULLIN1-RING ubiquitin ligase complexes (CRL1). The functionally redundant genes BLADE ON PETIOLE1 (BOP1) and -2 (BOP2) are potential candidates to regulate LFY activity and have recently been shown to be substrate adaptors of CULLIN3 (CUL3)-RING ubiquitin ligases (CRL3). We tested the hypothesis that LFY activity is controlled by BOPs and CUL3s in plants and that LFY is a substrate for ubiquitination by BOP-containing CRL3 complexes. When constitutively expressed, LFY activity is fully dependent on BOP2 as well as on CUL3A and B to regulate target genes such as APETALA1 and to induce ectopic flower formation. We also show that LFY and BOP2 proteins interact physically and that LFY-dependent ubiquitinated species are produced in vitro in a reconstituted cell-free CRL3 system in the presence of LFY, BOP2 and CUL3. This new post-translational regulation of LFY activity by CRL3 complexes makes it a unique transcription factor subjected to a positive dual regulation by both CRL1 and CRL3 complexes and suggests a novel mechanism for promoting flower development.
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http://dx.doi.org/10.1111/nph.15329DOI Listing
October 2018

Tetramerization of MADS family transcription factors SEPALLATA3 and AGAMOUS is required for floral meristem determinacy in Arabidopsis.

Nucleic Acids Res 2018 06;46(10):4966-4977

Laboratoire de Physiologie Cellulaire & Végétale, CEA, Univ. Grenoble Alpes, CNRS, INRA, BIG, Grenoble.

The MADS transcription factors (TF) constitute an ancient family of TF found in all eukaryotes that bind DNA as obligate dimers. Plants have dramatically expanded the functional diversity of the MADS family during evolution by adding protein-protein interaction domains to the core DNA-binding domain, allowing the formation of heterotetrameric complexes. Tetramerization of plant MADS TFs is believed to play a central role in the evolution of higher plants by acting as one of the main determinants of flower formation and floral organ specification. The MADS TF, SEPALLATA3 (SEP3), functions as a central protein-protein interaction hub, driving tetramerization with other MADS TFs. Here, we use a SEP3 splice variant, SEP3Δtet, which has dramatically abrogated tetramerization capacity to decouple SEP3 tetramerization and DNA-binding activities. We unexpectedly demonstrate that SEP3 heterotetramer formation is required for correct termination of the floral meristem, but plays a lesser role in floral organogenesis. The heterotetramer formed by SEP3 and the MADS protein, AGAMOUS, is necessary to activate two target genes, KNUCKLES and CRABSCLAW, which are required for meristem determinacy. These studies reveal unique and highly specific roles of tetramerization in flower development and suggest tetramerization may be required to activate only a subset of target genes in closed chromatin regions.
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http://dx.doi.org/10.1093/nar/gky205DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6007258PMC
June 2018

JASPAR 2018: update of the open-access database of transcription factor binding profiles and its web framework.

Nucleic Acids Res 2018 01;46(D1):D260-D266

Centre for Molecular Medicine Norway (NCMM), Nordic EMBL Partnership, University of Oslo, 0318 Oslo, Norway.

JASPAR (http://jaspar.genereg.net) is an open-access database of curated, non-redundant transcription factor (TF)-binding profiles stored as position frequency matrices (PFMs) and TF flexible models (TFFMs) for TFs across multiple species in six taxonomic groups. In the 2018 release of JASPAR, the CORE collection has been expanded with 322 new PFMs (60 for vertebrates and 262 for plants) and 33 PFMs were updated (24 for vertebrates, 8 for plants and 1 for insects). These new profiles represent a 30% expansion compared to the 2016 release. In addition, we have introduced 316 TFFMs (95 for vertebrates, 218 for plants and 3 for insects). This release incorporates clusters of similar PFMs in each taxon and each TF class per taxon. The JASPAR 2018 CORE vertebrate collection of PFMs was used to predict TF-binding sites in the human genome. The predictions are made available to the scientific community through a UCSC Genome Browser track data hub. Finally, this update comes with a new web framework with an interactive and responsive user-interface, along with new features. All the underlying data can be retrieved programmatically using a RESTful API and through the JASPAR 2018 R/Bioconductor package.
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http://dx.doi.org/10.1093/nar/gkx1126DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5753243PMC
January 2018

Structure of the TOPLESS corepressor provides insight into the evolution of transcriptional repression.

Proc Natl Acad Sci U S A 2017 07 11;114(30):8107-8112. Epub 2017 Jul 11.

Laboratoire de Physiologie Cellulaire et Végétale, Université Grenoble Alpes, CNRS, Commissariat à l'Energie Atomique et aux Energies Alternatives/Biosciences and Biotechnology Institute of Grenoble, Institut National de la Recherche Agronomique (INRA), F-38000 Grenoble, France.

Transcriptional repression involves a class of proteins called corepressors that link transcription factors to chromatin remodeling complexes. In plants such as , the most prominent corepressor is TOPLESS (TPL), which plays a key role in hormone signaling and development. Here we present the crystallographic structure of the TPL N-terminal region comprising the LisH and CTLH (C-terminal to LisH) domains and a newly identified third region, which corresponds to a CRA domain. Comparing the structure of TPL with the mammalian TBL1, which shares a similar domain structure and performs a parallel corepressor function, revealed that the plant TPLs have evolved a new tetramerization interface and unique and highly conserved surface for interaction with repressors. Using site-directed mutagenesis, we validated those surfaces in vitro and in vivo and showed that TPL tetramerization and repressor binding are interdependent. Our results illustrate how evolution used a common set of protein domains to create a diversity of corepressors, achieving similar properties with different molecular solutions.
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http://dx.doi.org/10.1073/pnas.1703054114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5544296PMC
July 2017

Plant SAM-Domain Proteins Start to Reveal Their Roles.

Trends Plant Sci 2017 08 28;22(8):718-725. Epub 2017 Jun 28.

LPCV, CEA, CNRS, INRA, Université Grenoble-Alpes, BIG, 38000 Grenoble, France. Electronic address:

Proteins often act in complexes assembled via protein-protein interaction domains. The sterile alpha motif (SAM) domain is one of the most prominent interaction domains in animals and is present in proteins of diverse functions. This domain allows head-to-tail closed oligomerisation or polymer formation resulting in homo- and/or heterocomplexes that have been shown to be important for proper protein localisation and function. In plants this domain is also present but has been poorly studied except for recent studies on the LEAFY floral regulator and the tRNA import component (TRIC)1/2 proteins. Here we catalogue SAM domain-containing proteins from arabidopsis (Arabidopsis thaliana), compare plant and other eukaryotic SAM domains, and perform homology modelling to probe plant SAM domain interaction capabilities.
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http://dx.doi.org/10.1016/j.tplants.2017.06.006DOI Listing
August 2017

A link between LEAFY and B-gene homologues in Welwitschia mirabilis sheds light on ancestral mechanisms prefiguring floral development.

New Phytol 2017 Oct 24;216(2):469-481. Epub 2017 Feb 24.

LPCV, CEA, CNRS, INRA, Université Grenoble-Alpes, BIG, 38000, Grenoble, France.

Flowering plants evolved from an unidentified gymnosperm ancestor. Comparison of the mechanisms controlling development in angiosperm flowers and gymnosperm cones may help to elucidate the mysterious origin of the flower. We combined gene expression studies with protein behaviour characterization in Welwitschia mirabilis to test whether the known regulatory links between LEAFY and its MADS-box gene targets, central to flower development, might also contribute to gymnosperm reproductive development. We found that WelLFY, one of two LEAFY-like genes in Welwitschia, could be an upstream regulator of the MADS-box genes APETALA3/PISTILLATA-like (B-genes). We demonstrated that, even though their DNA-binding domains are extremely similar, WelLFY and its paralogue WelNDLY exhibit distinct DNA-binding specificities, and that, unlike WelNDLY, WelLFY shares with its angiosperm orthologue the capacity to bind promoters of Welwitschia B-genes. Finally, we identified several cis-elements mediating these interactions in Welwitschia and obtained evidence that the link between LFY homologues and B-genes is also conserved in two other gymnosperms, Pinus and Picea. Although functional approaches to investigate cone development in gymnosperms are limited, our state-of-the-art biophysical techniques, coupled with expression studies, provide evidence that crucial links, central to the control of floral development, may already have existed before the appearance of flowers.
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http://dx.doi.org/10.1111/nph.14483DOI Listing
October 2017

Conservation vs divergence in LEAFY and APETALA1 functions between Arabidopsis thaliana and Cardamine hirsuta.

New Phytol 2017 Oct 18;216(2):549-561. Epub 2017 Jan 18.

Max Planck Institute for Plant Breeding Research, Carl-von-Linné-Weg 10, 50829, Köln, Germany.

A conserved genetic toolkit underlies the development of diverse floral forms among angiosperms. However, the degree of conservation vs divergence in the configuration of these gene regulatory networks is less clear. We addressed this question in a parallel genetic study between the closely related species Arabidopsis thaliana and Cardamine hirsuta. We identified leafy (lfy) and apetala1 (ap1) alleles in a mutant screen for floral regulators in C. hirsuta. C. hirsuta lfy mutants showed a complete homeotic conversion of flowers to leafy shoots, mimicking lfy ap1 double mutants in A. thaliana. Through genetic and molecular experiments, we showed that AP1 activation is fully dependent on LFY in C. hirsuta, by contrast to A. thaliana. Additionally, we found that LFY influences heteroblasty in C. hirsuta, such that loss or gain of LFY function affects its progression. Overexpression of UNUSUAL FLORAL ORGANS also alters C. hirsuta leaf shape in an LFY-dependent manner. We found that LFY and AP1 are conserved floral regulators that act nonredundantly in C. hirsuta, such that LFY has more obvious roles in floral and leaf development in C. hirsuta than in A. thaliana.
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http://dx.doi.org/10.1111/nph.14419DOI Listing
October 2017

A flower is born: an update on Arabidopsis floral meristem formation.

Curr Opin Plant Biol 2017 02 6;35:15-22. Epub 2016 Oct 6.

Laboratory of Plant & Cell Physiology, CNRS, CEA, Univ. Grenoble Alpes, INRA, 38000 Grenoble, France. Electronic address:

In Arabidopsis, floral meristems appear on the flanks of the inflorescence meristem. Their stereotypic development, ultimately producing the four whorls of floral organs, is essentially controlled by a network coordinating growth and cell-fate determination. This network integrates hormonal signals, transcriptional regulators, and mechanical constraints. Mechanisms regulating floral meristem formation have been studied at many different scales, from protein structure to tissue modeling. In this paper, we review recent findings related to the emergence of the floral meristem and floral fate determination and examine how this field has been impacted by recent technological developments.
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http://dx.doi.org/10.1016/j.pbi.2016.09.003DOI Listing
February 2017

Deciphering the Molecular Mechanisms Underpinning the Transcriptional Control of Gene Expression by Master Transcriptional Regulators in Arabidopsis Seed.

Plant Physiol 2016 06 12;171(2):1099-112. Epub 2016 Apr 12.

Institut Jean-Pierre Bourgin, Institut National de la Recherche Agronomique, AgroParisTech, Centre National de la Recherche Scientifique, Université Paris-Saclay, RD10, F-78026 Versailles cedex, France (S.Ba., Z.K., J.T., C.B., A.T., M.P., N.B., D.E.-C., L.L., B.D.);Laboratoire de Physiologie Cellulaire et Végétale, Université Grenoble Alpes, Centre National de la Recherche Scientifique Unité Mixte de Recherche 5168, Commissariat à l'Energie Atomique/DRF/BIG, Institut National de la Recherche Agronomique Unité Mixte de Recherche 1417, 38054 Grenoble, France (S.Bl., E.T., F.P.); andGenomics Unit (J.M.F.-Z., M.G.) and Plant Molecular Genetics Department (R.S.), Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Campus Universidad Autónoma, 28049 Madrid, Spain

In Arabidopsis (Arabidopsis thaliana), transcriptional control of seed maturation involves three related regulators with a B3 domain, namely LEAFY COTYLEDON2 (LEC2), ABSCISIC ACID INSENSITIVE3 (ABI3), and FUSCA3 (ABI3/FUS3/LEC2 [AFLs]). Although genetic analyses have demonstrated partially overlapping functions of these regulators, the underlying molecular mechanisms remained elusive. The results presented here confirmed that the three proteins bind RY DNA elements (with a 5'-CATG-3' core sequence) but with different specificities for flanking nucleotides. In planta as in the moss Physcomitrella patens protoplasts, the presence of RY-like (RYL) elements is necessary but not sufficient for the regulation of the OLEOSIN1 (OLE1) promoter by the B3 AFLs. G box-like domains, located in the vicinity of the RYL elements, also are required for proper activation of the promoter, suggesting that several proteins are involved. Consistent with this idea, LEC2 and ABI3 showed synergistic effects on the activation of the OLE1 promoter. What is more, LEC1 (a homolog of the NF-YB subunit of the CCAAT-binding complex) further enhanced the activation of this target promoter in the presence of LEC2 and ABI3. Finally, recombinant LEC1 and LEC2 proteins produced in Arabidopsis protoplasts could form a ternary complex with NF-YC2 in vitro, providing a molecular explanation for their functional interactions. Taken together, these results allow us to propose a molecular model for the transcriptional regulation of seed genes by the L-AFL proteins, based on the formation of regulatory multiprotein complexes between NF-YBs, which carry a specific aspartate-55 residue, and B3 transcription factors.
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http://dx.doi.org/10.1104/pp.16.00034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4902591PMC
June 2016

A SAM oligomerization domain shapes the genomic binding landscape of the LEAFY transcription factor.

Nat Commun 2016 Apr 21;7:11222. Epub 2016 Apr 21.

Laboratoire de Physiologie Cellulaire et Végétale, Université Grenoble Alpes, CNRS UMR5168, CEA/DRF/BIG, INRA UMR 1417, 17, avenue des Martyrs, 38054 Grenoble, France.

Deciphering the mechanisms directing transcription factors (TFs) to specific genome regions is essential to understand and predict transcriptional regulation. TFs recognize short DNA motifs primarily through their DNA-binding domain. Some TFs also possess an oligomerization domain suspected to potentiate DNA binding but for which the genome-wide influence remains poorly understood. Here we focus on the LEAFY transcription factor, a master regulator of flower development in angiosperms. We have determined the crystal structure of its conserved amino-terminal domain, revealing an unanticipated Sterile Alpha Motif oligomerization domain. We show that this domain is essential to LEAFY floral function. Moreover, combined biochemical and genome-wide assays suggest that oligomerization is required for LEAFY to access regions with low-affinity binding sites or closed chromatin. This finding shows that domains that do not directly contact DNA can nevertheless have a profound impact on the DNA binding landscape of a TF.
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http://dx.doi.org/10.1038/ncomms11222DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4844672PMC
April 2016

A Glimpse beyond Structures in Auxin-Dependent Transcription.

Trends Plant Sci 2016 07 22;21(7):574-583. Epub 2016 Mar 22.

Laboratoire de Physiologie Cellulaire et Végétale, Université Grenoble Alpes, CNRS UMR5168, CEA/DRF/BIG, INRA UMR 1417, Grenoble, France.

Auxin response factors (ARFs), transcription factors (TFs), and their Aux/IAA (IAA) repressors are central components of the auxin signalling pathway. They interact as homo- and heteromultimers. The structure of their interacting domains revealed a PB1 fold mediating electrostatic interactions through positive and negative faces. Detailed structural analysis revealed additional hydrophobic and polar determinants and started unveiling an ARF/IAA interaction code. Structural progress also shed new light on the DNA binding mode of ARFs showing how they dimerize to bind repeated DNA elements. Here, we discuss the in vitro and in vivo significance of these structural properties for the ARF family of TFs and identify some critical missing information on how specificity might be achieved in the auxin signalling pathway.
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http://dx.doi.org/10.1016/j.tplants.2016.02.002DOI Listing
July 2016

The Myb-domain protein ULTRAPETALA1 INTERACTING FACTOR 1 controls floral meristem activities in Arabidopsis.

Development 2016 Apr 22;143(7):1108-19. Epub 2016 Feb 22.

Université Grenoble Alpes, Laboratoire de Physiologie Cellulaire et Végétale (LPCV), Grenoble 38054, France CNRS, LPCV, UMR 5168, Grenoble 38054, France CEA, Direction des Sciences du Vivant, BIG, LPCV, Grenoble 38054, France INRA, LPCV, Grenoble 38054, France

Higher plants continuously and iteratively produce new above-ground organs in the form of leaves, stems and flowers. These organs arise from shoot apical meristems whose homeostasis depends on coordination between self-renewal of stem cells and their differentiation into organ founder cells. This coordination is stringently controlled by the central transcription factor WUSCHEL (WUS), which is both necessary and sufficient for stem cell specification in Arabidopsis thaliana ULTRAPETALA1 (ULT1) was previously identified as a plant-specific, negative regulator of WUS expression. However, molecular mechanisms underlying this regulation remain unknown. ULT1 protein contains a SAND putative DNA-binding domain and a B-box, previously proposed as a protein interaction domain in eukaryotes. Here, we characterise a novel partner of ULT1, named ULT1 INTERACTING FACTOR 1 (UIF1), which contains a Myb domain and an EAR motif. UIF1 and ULT1 function in the same pathway for regulation of organ number in the flower. Moreover, UIF1 displays DNA-binding activity and specifically binds to WUS regulatory elements. We thus provide genetic and molecular evidence that UIF1 and ULT1 work together in floral meristem homeostasis, probably by direct repression of WUS expression.
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http://dx.doi.org/10.1242/dev.127365DOI Listing
April 2016

Evolution of the Plant Reproduction Master Regulators LFY and the MADS Transcription Factors: The Role of Protein Structure in the Evolutionary Development of the Flower.

Front Plant Sci 2015 6;6:1193. Epub 2016 Jan 6.

CNRS, Laboratoire de Physiologie Cellulaire & Végétale, UMR 5168Grenoble, France; Laboratoire de Physiologie Cellulaire & Végétale, University of Grenoble AlpesGrenoble, France; Commissariat à l´Energie Atomique et aux Energies Alternatives, Direction des Sciences du Vivant, Laboratoire de Physiologie Cellulaire & Végétale, Institut de Recherches en Technologies et Sciences pour le VivantGrenoble, France; Laboratoire de Physiologie Cellulaire & Végétale, Institut National de la Recherche AgronomiqueGrenoble, France.

Understanding the evolutionary leap from non-flowering (gymnosperms) to flowering (angiosperms) plants and the origin and vast diversification of the floral form has been one of the focuses of plant evolutionary developmental biology. The evolving diversity and increasing complexity of organisms is often due to relatively small changes in genes that direct development. These "developmental control genes" and the transcription factors (TFs) they encode, are at the origin of most morphological changes. TFs such as LEAFY (LFY) and the MADS-domain TFs act as central regulators in key developmental processes of plant reproduction including the floral transition in angiosperms and the specification of the male and female organs in both gymnosperms and angiosperms. In addition to advances in genome wide profiling and forward and reverse genetic screening, structural techniques are becoming important tools in unraveling TF function by providing atomic and molecular level information that was lacking in purely genetic approaches. Here, we summarize previous structural work and present additional biophysical and biochemical studies of the key master regulators of plant reproduction - LEAFY and the MADS-domain TFs SEPALLATA3 and AGAMOUS. We discuss the impact of structural biology on our understanding of the complex evolutionary process leading to the development of the bisexual flower.
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http://dx.doi.org/10.3389/fpls.2015.01193DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4701952PMC
January 2016

JASPAR 2016: a major expansion and update of the open-access database of transcription factor binding profiles.

Nucleic Acids Res 2016 Jan 3;44(D1):D110-5. Epub 2015 Nov 3.

Centre for Molecular Medicine and Therapeutics at the Child and Family Research Institute, Department of Medical Genetics, University of British Columbia, Vancouver, V5Z 4H4, BC, Canada

JASPAR (http://jaspar.genereg.net) is an open-access database storing curated, non-redundant transcription factor (TF) binding profiles representing transcription factor binding preferences as position frequency matrices for multiple species in six taxonomic groups. For this 2016 release, we expanded the JASPAR CORE collection with 494 new TF binding profiles (315 in vertebrates, 11 in nematodes, 3 in insects, 1 in fungi and 164 in plants) and updated 59 profiles (58 in vertebrates and 1 in fungi). The introduced profiles represent an 83% expansion and 10% update when compared to the previous release. We updated the structural annotation of the TF DNA binding domains (DBDs) following a published hierarchical structural classification. In addition, we introduced 130 transcription factor flexible models trained on ChIP-seq data for vertebrates, which capture dinucleotide dependencies within TF binding sites. This new JASPAR release is accompanied by a new web tool to infer JASPAR TF binding profiles recognized by a given TF protein sequence. Moreover, we provide the users with a Ruby module complementing the JASPAR API to ease programmatic access and use of the JASPAR collection of profiles. Finally, we provide the JASPAR2016 R/Bioconductor data package with the data of this release.
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http://dx.doi.org/10.1093/nar/gkv1176DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4702842PMC
January 2016

MORPHEUS, a Webtool for Transcription Factor Binding Analysis Using Position Weight Matrices with Dependency.

PLoS One 2015 18;10(8):e0135586. Epub 2015 Aug 18.

Laboratoire de Physiologie Cellulaire Végétale, Unité Mixte de Recherche 5168, Centre National de la Recherche Scientifique, Commissariat à l'Énergie Atomique, Institut National de la Recherche Agronomique, Université Joseph Fourier Grenoble I, 38054, Grenoble, France.

Transcriptional networks are central to any biological process and changes affecting transcription factors or their binding sites in the genome are a key factor driving evolution. As more organisms are being sequenced, tools are needed to easily predict transcription factor binding sites (TFBS) presence and affinity from mere inspection of genomic sequences. Although many TFBS discovery algorithms exist, tools for using the DNA binding models they generate are relatively scarce and their use is limited among the biologist community by the lack of flexible and user-friendly tools. We have developed a suite of web tools (called Morpheus) based on the proven Position Weight Matrices (PWM) formalism that can be used without any programing skills and incorporates some unique features such as the presence of dependencies between nucleotides positions or the possibility to compute the predicted occupancy of a large regulatory region using a biophysical model. To illustrate the possibilities and simplicity of Morpheus tools in functional and evolutionary analysis, we have analysed the regulatory link between LEAFY, a key plant transcription factor involved in flower development, and its direct target gene APETALA1 during the divergence of Brassicales clade.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0135586PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4540572PMC
May 2016

Evolution. Response to Comment on "A promiscuous intermediate underlies the evolution of LEAFY DNA binding specificity".

Science 2015 Feb;347(6222):621

CNRS, Laboratoire de Physiologie Cellulaire et Végétale (LPCV), UMR 5168, 38054 Grenoble, France. UGA, LPCV, F-38054 Grenoble, France. Commissariat à l'énergie atomique et aux énergies alternatives, Direction des Sciences du Vivant, Institut de Recherches en Technologies et Sciences pour le Vivant, LPCV, F-38054 Grenoble, France. Institut National de la Recherche Agronomique, LPCV, F-38054 Grenoble, France.

Brunkard et al. propose that the identification of novel LEAFY sequences contradicts our model of evolution through promiscuous intermediates. Based on the debate surrounding land plant phylogeny and on our analysis of these interesting novel sequences, we explain why there is no solid evidence to disprove our model.
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http://dx.doi.org/10.1126/science.1256011DOI Listing
February 2015

Structural basis for the oligomerization of the MADS domain transcription factor SEPALLATA3 in Arabidopsis.

Plant Cell 2014 Sep 16;26(9):3603-15. Epub 2014 Sep 16.

CNRS, Laboratoire de Physiologie Cellulaire and Végétale, UMR 5168, 38054 Grenoble, France Université Grenoble Alpes, Laboratoire de Physiologie Cellulaire et Végétale, F-38054 Grenoble, France Commissariat à l'Energie Atomique, Direction des Sciences du Vivant, Institut de Recherches en Technologies et Sciences pour le Vivant, Laboratoire de Physiologie Cellulaire et Végétale, F-38054 Grenoble, France INRA, Laboratoire de Physiologie Cellulaire et Végétale, USC1359, F-38054 Grenoble, France

In plants, MADS domain transcription factors act as central regulators of diverse developmental pathways. In Arabidopsis thaliana, one of the most central members of this family is SEPALLATA3 (SEP3), which is involved in many aspects of plant reproduction, including floral meristem and floral organ development. SEP3 has been shown to form homo and heterooligomeric complexes with other MADS domain transcription factors through its intervening (I) and keratin-like (K) domains. SEP3 function depends on its ability to form specific protein-protein complexes; however, the atomic level determinants of oligomerization are poorly understood. Here, we report the 2.5-Å crystal structure of a small portion of the intervening and the complete keratin-like domain of SEP3. The domains form two amphipathic alpha helices separated by a rigid kink, which prevents intramolecular association and presents separate dimerization and tetramerization interfaces comprising predominantly hydrophobic patches. Mutations to the tetramerization interface demonstrate the importance of highly conserved hydrophobic residues for tetramer stability. Atomic force microscopy was used to show SEP3-DNA interactions and the role of oligomerization in DNA binding and conformation. Based on these data, the oligomerization patterns of the larger family of MADS domain transcription factors can be predicted and manipulated based on the primary sequence.
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http://dx.doi.org/10.1105/tpc.114.127910DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4213154PMC
September 2014

Structural basis for oligomerization of auxin transcriptional regulators.

Nat Commun 2014 Apr 7;5:3617. Epub 2014 Apr 7.

1] CNRS, Laboratoire de Physiologie Cellulaire and Végétale, UMR 5168, Grenoble 38054, France [2] Univ. Grenoble Alpes, LPCV, Grenoble F-38054, France [3] CEA, DSV, iRTSV, LPCV, Grenoble F-38054, France [4] INRA, LPCV, Grenoble F-38054, France.

The plant hormone auxin is a key morphogenetic regulator acting from embryogenesis onwards. Transcriptional events in response to auxin are mediated by the auxin response factor (ARF) transcription factors and the Aux/IAA (IAA) transcriptional repressors. At low auxin concentrations, IAA repressors associate with ARF proteins and recruit corepressors that prevent auxin-induced gene expression. At higher auxin concentrations, IAAs are degraded and ARFs become free to regulate auxin-responsive genes. The interaction between ARFs and IAAs is thus central to auxin signalling and occurs through the highly conserved domain III/IV present in both types of proteins. Here, we report the crystal structure of ARF5 domain III/IV and reveal the molecular determinants of ARF-IAA interactions. We further provide evidence that ARFs have the potential to oligomerize, a property that could be important for gene regulation in response to auxin.
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http://dx.doi.org/10.1038/ncomms4617DOI Listing
April 2014