An early secretory pathway mediated by GNOM-LIKE 1 and GNOM is essential for basal polarity establishment in Arabidopsis thaliana

Spatial regulation of the plant hormone indole-3-acetic acid (IAA, or auxin) is essential for plant development. Auxin gradient establishment is mediated by polarly localized auxin transporters, including PIN-FORMED (PIN) proteins. Their localization and abundance at the plasma membrane are tightly regulated by endo-membrane machinery, especially the endocytic and recycling pathways mediated by the ADP ribosylation factor guanine nucleotide exchange factor (ARF-GEF) GNOM. We assessed the role of the early secretory pathway in establishing PIN1 polarity in Arabidopsis thaliana by pharmacological and genetic approaches. We identified the compound endosidin 8 (ES8), which selectively interferes with PIN1 basal polarity without altering the polarity of apical proteins. ES8 alters the auxin distribution pattern in the root and induces a strong developmental phenotype, including reduced root length. The ARF-GEF-defective mutants gnom-like 1 (gnl1-1) and gnom (van7) are significantly resistant to ES8. The compound does not affect recycling or vacuolar trafficking of PIN1 but leads to its intracellular accumulation, resulting in loss of PIN1 basal polarity at the plasma membrane. Our data confirm a role for GNOM in endoplasmic reticulum (ER)-Golgi trafficking and reveal that a GNL1/GNOM-mediated early secretory pathway selectively regulates PIN1 basal polarity establishment in a manner essential for normal plant development

A network of stress-related genes regulates hypocotyl elongation downstream of selective auxin perception

The plant hormone auxin, a master coordinator of development, regulates hypocotyl elongation during seedling growth. We previously identified the synthetic molecule RubNeddin 1 (RN1), which induces degradation of the AUXIN/INDOLE-3-ACETIC ACID (AUX/IAA) transcriptional repressors INDOLE-3-ACETIC ACID-INDUCIBLE3 (IAA3) and IAA7 in planta and strongly promotes hypocotyl elongation. In the present study, we show that despite the structural similarity of RN1 to the synthetic auxin 2,4-dichlorophenoxyacetic-acid (2,4-D), direct treatments with these compounds in Arabidopsis (Arabidopsis thaliana) result in distinct effects, possibly due to enhanced uptake of RN1 and low-level, chronic release of 2,4-D from RN1 in planta. We confirm RN1-induced hypocotyl elongation occurs via specific TRANSPORT INHIBITOR RESISTANT1 (TIR1)/AUXIN SIGNALING F-BOX (AFB) receptor-mediated auxin signaling involving TIR1, AFB2, and AFB5. Using a transcriptome profiling strategy and candidate gene approach, we identify the genes ZINC FINGER OF ARABIDOPSIS THALIANA10 (ZAT10), ARABIDOPSIS TOXICOS EN LEVADURA31 (ATL31), and WRKY DNA-BINDING PROTEIN33 (WRKY33) as being rapidly upregulated by RN1, despite being downregulated by 2,4-D treatment. RN1-induced expression of these genes also occurs via TIR1/AFB-mediated auxin signaling. Our results suggest both hypocotyl elongation and transcription of these genes are induced by RN1 via the promoted degradation of the AUX/IAA transcriptional repressor IAA7. Moreover, these three genes, which are known to be stress-related, act in an inter-dependent transcriptional regulatory network controlling hypocotyl elongation. Together, our results suggest ZAT10, ATL31, and WRKY33 take part in a common gene network regulating hypocotyl elongation in Arabidopsis downstream of a selective auxin perception module likely involving TIR1, AFB2, and AFB5 and inducing the degradation of IAA7

The AINTEGUMENTA LIKE1

Adventitious rooting is an essential but sometimes rate-limiting step in the clonal multiplication of elite tree germplasm, because the ability to form roots declines rapidly with age in mature adult plant tissues. In spite of the importance of adventitious rooting, the mechanism behind this developmental process remains poorly understood. We have described the transcriptional profiles that are associated with the developmental stages of adventitious root formation in the model tree poplar (Populus trichocarpa). Transcriptome analyses indicate a highly specific temporal induction of the AINTEGUMENTA LIKE1 (PtAIL1) transcription factor of the AP2 family during adventitious root formation. Transgenic poplar samples that overexpressed PtAIL1 were able to grow an increased number of adventitious roots, whereas RNA interference mediated the down-expression of PtAIL1 expression, which led to a delay in adventitious root formation. Microarray analysis showed that the expression of 15 genes, including the transcription factors AGAMOUS-Like6 and MYB36, was overexpressed in the stem tissues that generated root primordia in PtAIL1-overexpressing plants, whereas their expression was reduced in the RNA interference lines. These results demonstrate that PtAIL1 is a positive regulator of poplar rooting that acts early in the development of adventitious roots

Selective auxin agonists induce specific AUX/IAA protein degradation to modulate plant development.

Auxin phytohormones control most aspects of plant development through a complex and interconnected signaling network. In the presence of auxin, AUXIN/INDOLE-3-ACETIC ACID (AUX/IAA) transcriptional repressors are targeted for degradation by the SKP1-CULLIN1-F-BOX (SCF) ubiquitin-protein ligases containing TRANSPORT INHIBITOR RESISTANT 1/AUXIN SIGNALING F-BOX (TIR1/AFB). CULLIN1-neddylation is required for SCFTIR1/AFB functionality, as exemplified by mutants deficient in the NEDD8-activating enzyme subunit AUXIN-RESISTANT 1 (AXR1). Here, we report a chemical biology screen that identifies small molecules requiring AXR1 to modulate plant development. We selected four molecules of interest, RubNeddin 1 to 4 (RN1 to -4), among which RN3 and RN4 trigger selective auxin responses at transcriptional, biochemical, and morphological levels. This selective activity is explained by their ability to consistently promote the interaction between TIR1 and a specific subset of AUX/IAA proteins, stimulating the degradation of particular AUX/IAA combinations. Finally, we performed a genetic screen using RN4, the RN with the greatest potential for dissecting auxin perception, which revealed that the chromatin remodeling ATPase BRAHMA is implicated in auxin-mediated apical hook development. These results demonstrate the power of selective auxin agonists to dissect auxin perception for plant developmental functions, as well as offering opportunities to discover new molecular players involved in auxin responses

The CEP5 Peptide Promotes Abiotic Stress Tolerance, As Revealed by Quantitative Proteomics, and Attenuates the AUX/IAA Equilibrium in Arabidopsis.

Peptides derived from non-functional precursors play important roles in various developmental processes, but also in (a)biotic stress signaling. Our (phospho)proteome-wide analyses of C-TERMINALLY ENCODED PEPTIDE 5 (CEP5)-mediated changes revealed an impact on abiotic stress-related processes. Drought has a dramatic impact on plant growth, development and reproduction, and the plant hormone auxin plays a role in drought responses. Our genetic, physiological, biochemical, and pharmacological results demonstrated that CEP5-mediated signaling is relevant for osmotic and drought stress tolerance in Arabidopsis, and that CEP5 specifically counteracts auxin effects. Specifically, we found that CEP5 signaling stabilizes AUX/IAA transcriptional repressors, suggesting the existence of a novel peptide-dependent control mechanism that tunes auxin signaling. These observations align with the recently described role of AUX/IAAs in stress tolerance and provide a novel role for CEP5 in osmotic and drought stress tolerance

Analysis of molecular mechanisms involved in adventitious root formation in poplar : The role of the PtAIL1 transcription factor

Ma thèse s'inscrit dans le cadre du projet européen ENERGYPOPLAR, dont l'objectif est d'améliorer la production de biomasse d'hybrides de Peuplier dans un système de sylviculture intensive. La propagation clonale, utilisant des boutures, est un outil approprié dans la production rapide. Une étape clé de cette propagation clonale est la formation de racines adventives. Ainsi, j'ai analysé la capacité à enraciner de plusieurs génotypes de peuplier sélectionnés. Cette étude suggère que certains génotypes présentent des facilités à enraciner par rapport à d'autres. Le principal objectif de la thèse a été d'identifier des gènes clefs de la rhizogenèse adventive du peuplier, plus précisément le rôle des facteurs de transcription (FT). J'ai débuté cette étude, par une analyse transcriptomique globale à différents stades de la formation de la racine adventive de P. trichocarpa. Durant le développement de la racine adventive, des transcrits des familles PIN, GH3, AUX/IAA, AUX/LAX et ARF sont régulés suggérant l'implication des voies de signalisation de l'auxine. De plus, cette analyse a révélé la régulation de l'expression de la famille de facteurs de transcription AP2/ERF, particulièrement le membre AIL1. Des études phénotypiques des lignées transgéniques de Peuplier (35S::AIL1 et RNAi), indiquent une variation dans la formation et dans le nombre de racines adventives. Dans le but de décrypter le réseau de gènes associé à l'expression de PtAIL1, j'ai réalisé une analyse transcriptomique des plantes transgéniques et du sauvage. Cette analyse a permis de suggérer des cibles potentielles de PtAIL1. Mes résultats montrent le rôle de PtAIL1 dans la formation des racines adventivesMy thesis was part of the european project ENERGYPOPLAR aiming to improve biomass yield of poplar hybrids in a intensive forestry system. Clonal propagation, by use of cuttings, is a very powerful tool in the fast production of poplars. A key step of this process is the adventitious root formation. Therefore, I investigated the rooting behavior of several selected poplar genotypes. Differences in rooting capacity could be shown suggesting the existence of easy-to-root and difficult-to-root genotypes. The main goal of my thesis was to identify key genes involved in the adventitious rooting process in poplar, with particular interest in the role of transcription factors (TFs). An initial genome-wide transcript profiling of the different stages of adventitious root formation in P. trichocarpa was carried-out. Auxin signaling seems to be involved in poplar adventitious root development, since the transcript level of several gene families like PIN, GH3, AUX/IAA, AUX/LAX and ARF were regulated. Further, the AP2/ERF transcription factors family and in particular AIL1 were identified as important factors involved in adventitious rooting. The phenotypic characterization of poplar transgenics lines (35s::AIL1 and RNAi lines) showed differences in the chronology of adventitious root formation and in the number of roots formed. In order to investigate the pathway(s) associated with PtAIL1 expression, transcriptomic analyses of transgenic lines and wild type were performed. Agamous-like and myb transcription factors were identified as possible candidate genes.PtAIL1 is one of the key regulators involved in adventitious root formation in popla

Etude des mécanismes moléculaires impliqués dans la formation des racines adventives du peuplier : rôle du facteur de transcription PtAIL1

My thesis was part of the european project ENERGYPOPLAR aiming to improve biomass yield of poplar hybrids in a intensive forestry system. Clonal propagation, by use of cuttings, is a very powerful tool in the fast production of poplars. A key step of this process is the adventitious root formation. Therefore, I investigated the rooting behavior of several selected poplar genotypes. Differences in rooting capacity could be shown suggesting the existence of easy-to-root and difficult-to-root genotypes. The main goal of my thesis was to identify key genes involved in the adventitious rooting process in poplar, with particular interest in the role of transcription factors (TFs). An initial genome-wide transcript profiling of the different stages of adventitious root formation in P. trichocarpa was carried-out. Auxin signaling seems to be involved in poplar adventitious root development, since the transcript level of several gene families like PIN, GH3, AUX/IAA, AUX/LAX and ARF were regulated. Further, the AP2/ERF transcription factors family and in particular AIL1 were identified as important factors involved in adventitious rooting. The phenotypic characterization of poplar transgenics lines (35s::AIL1 and RNAi lines) showed differences in the chronology of adventitious root formation and in the number of roots formed. In order to investigate the pathway(s) associated with PtAIL1 expression, transcriptomic analyses of transgenic lines and wild type were performed. Agamous-like and myb transcription factors were identified as possible candidate genes.PtAIL1 is one of the key regulators involved in adventitious root formation in poplarMa thèse s'inscrit dans le cadre du projet européen ENERGYPOPLAR, dont l'objectif est d'améliorer la production de biomasse d'hybrides de Peuplier dans un système de sylviculture intensive. La propagation clonale, utilisant des boutures, est un outil approprié dans la production rapide. Une étape clé de cette propagation clonale est la formation de racines adventives. Ainsi, j'ai analysé la capacité à enraciner de plusieurs génotypes de peuplier sélectionnés. Cette étude suggère que certains génotypes présentent des facilités à enraciner par rapport à d'autres. Le principal objectif de la thèse a été d'identifier des gènes clefs de la rhizogenèse adventive du peuplier, plus précisément le rôle des facteurs de transcription (FT). J'ai débuté cette étude, par une analyse transcriptomique globale à différents stades de la formation de la racine adventive de P. trichocarpa. Durant le développement de la racine adventive, des transcrits des familles PIN, GH3, AUX/IAA, AUX/LAX et ARF sont régulés suggérant l'implication des voies de signalisation de l'auxine. De plus, cette analyse a révélé la régulation de l'expression de la famille de facteurs de transcription AP2/ERF, particulièrement le membre AIL1. Des études phénotypiques des lignées transgéniques de Peuplier (35S::AIL1 et RNAi), indiquent une variation dans la formation et dans le nombre de racines adventives. Dans le but de décrypter le réseau de gènes associé à l'expression de PtAIL1, j'ai réalisé une analyse transcriptomique des plantes transgéniques et du sauvage. Cette analyse a permis de suggérer des cibles potentielles de PtAIL1. Mes résultats montrent le rôle de PtAIL1 dans la formation des racines adventive

Etude des mécanismes moléculaires impliqués dans la formation des racines adventives du peuplier (rôle du facteur de transcription PtAIL1)

Ma thèse s'inscrit dans le cadre du projet européen ENERGYPOPLAR, dont l'objectif est d'améliorer la production de biomasse d'hybrides de Peuplier dans un système de sylviculture intensive. La propagation clonale, utilisant des boutures, est un outil approprié dans la production rapide. Une étape clé de cette propagation clonale est la formation de racines adventives. Ainsi, j'ai analysé la capacité à enraciner de plusieurs génotypes de peuplier sélectionnés. Cette étude suggère que certains génotypes présentent des facilités à enraciner par rapport à d'autres. Le principal objectif de la thèse a été d'identifier des gènes clefs de la rhizogenèse adventive du peuplier, plus précisément le rôle des facteurs de transcription (FT). J'ai débuté cette étude, par une analyse transcriptomique globale à différents stades de la formation de la racine adventive de P. trichocarpa. Durant le développement de la racine adventive, des transcrits des familles PIN, GH3, AUX/IAA, AUX/LAX et ARF sont régulés suggérant l'implication des voies de signalisation de l'auxine. De plus, cette analyse a révélé la régulation de l'expression de la famille de facteurs de transcription AP2/ERF, particulièrement le membre AIL1. Des études phénotypiques des lignées transgéniques de Peuplier (35S::AIL1 et RNAi), indiquent une variation dans la formation et dans le nombre de racines adventives. Dans le but de décrypter le réseau de gènes associé à l'expression de PtAIL1, j'ai réalisé une analyse transcriptomique des plantes transgéniques et du sauvage. Cette analyse a permis de suggérer des cibles potentielles de PtAIL1. Mes résultats montrent le rôle de PtAIL1 dans la formation des racines adventivesMy thesis was part of the european project ENERGYPOPLAR aiming to improve biomass yield of poplar hybrids in a intensive forestry system. Clonal propagation, by use of cuttings, is a very powerful tool in the fast production of poplars. A key step of this process is the adventitious root formation. Therefore, I investigated the rooting behavior of several selected poplar genotypes. Differences in rooting capacity could be shown suggesting the existence of easy-to-root and difficult-to-root genotypes. The main goal of my thesis was to identify key genes involved in the adventitious rooting process in poplar, with particular interest in the role of transcription factors (TFs). An initial genome-wide transcript profiling of the different stages of adventitious root formation in P. trichocarpa was carried-out. Auxin signaling seems to be involved in poplar adventitious root development, since the transcript level of several gene families like PIN, GH3, AUX/IAA, AUX/LAX and ARF were regulated. Further, the AP2/ERF transcription factors family and in particular AIL1 were identified as important factors involved in adventitious rooting. The phenotypic characterization of poplar transgenics lines (35s::AIL1 and RNAi lines) showed differences in the chronology of adventitious root formation and in the number of roots formed. In order to investigate the pathway(s) associated with PtAIL1 expression, transcriptomic analyses of transgenic lines and wild type were performed. Agamous-like and myb transcription factors were identified as possible candidate genes.PtAIL1 is one of the key regulators involved in adventitious root formation in poplarMETZ-SCD (574632105) / SudocNANCY1-Bib. numérique (543959902) / SudocNANCY2-Bibliotheque electronique (543959901) / SudocNANCY-INPL-Bib. électronique (545479901) / SudocSudocFranceF

The AINTEGUMENTA LIKE1 Homeotic Transcription Factor PtAIL1 Controls the Formation of Adventitious Root Primordia in Poplar

Adventitious rooting is an essential but sometimes rate-limiting step in the clonal multiplication of elite tree germplasm, because the ability to form roots declines rapidly with age in mature adult plant tissues. In spite of the importance of adventitious rooting, the mechanism behind this developmental process remains poorly understood. We have described the transcriptional profiles that are associated with the developmental stages of adventitious root formation in the model tree poplar (Populus trichocarpa). Transcriptome analyses indicate a highly specific temporal induction of the AINTEGUMENTA LIKE1 (PtAIL1) transcription factor of the AP2 family during adventitious root formation. Transgenic poplar samples that overexpressed PtAIL1 were able to grow an increased number of adventitious roots, whereas RNA interference mediated the down-expression of PtAIL1 expression, which led to a delay in adventitious root formation. Microarray analysis showed that the expression of 15 genes, including the transcription factors AGAMOUS-Like6 and MYB36, was overexpressed in the stem tissues that generated root primordia in PtAIL1-overexpressing plants, whereas their expression was reduced in the RNA interference lines. These results demonstrate that PtAIL1 is a positive regulator of poplar rooting that acts early in the development of adventitious roots

Molecular basis of differential adventitious rooting competence in poplar genotypes

Recalcitrant adventitious root (AR) development is a major hurdle in propagating commercially important woody plants. Although significant progress has been made to identify genes involved in subsequent steps of AR development, the molecular basis of differences in apparent recalcitrance to form AR between easy-to-root and difficult-to-root genotypes remains unknown. To address this, we generated cambium tissue-specific transcriptomic data from stem cuttings of hybrid aspen, T89 (difficult-to-root) and hybrid poplar OP42 (easy-to-root), and used transgenic approaches to verify the role of several transcription factors in the control of adventitious rooting. Increased peroxidase activity was positively correlated with better rooting. We found differentially expressed genes encoding reactive oxygen species scavenging proteins to be enriched in OP42 compared with T89. A greater number of differentially expressed transcription factors in cambium cells of OP42 compared with T89 was revealed by a more intense transcriptional reprograming in the former. PtMYC2, a potential negative regulator, was less expressed in OP42 compared with T89. Using transgenic approaches, we demonstrated that PttARF17.1 and PttMYC2.1 negatively regulate adventitious rooting. Our results provide insights into the molecular basis of genotypic differences in AR and implicate differential expression of the master regulator MYC2 as a critical player in this process