Letteratura scientifica selezionata sul tema "Root meristems"

We have previously described the phenotype of Arabidopsis thaliana plants with mutations at the CLAVATA1 (CLV1) locus (Clark, S. E., Running, M. P. and Meyerowitz, E. M. (1993) Development 119, 397–418). Our investigations demonstrated that clv1 plants develop enlarged vegetative and inflorescence apical meristems, and enlarged and indeterminate floral meristems. Here, we present an analysis of mutations at a separate locus, CLAVATA3 (CLV3), that disrupt meristem development in a manner similar to clv1 mutations. clv3 plants develop enlarged apical meristems as early as the mature embryo stage. clv3 floral meristems are also enlarged compared with wild type, and maintain a proliferating meristem throughout flower development. clv3 root meristems are unaffected, indicating that CLV3 is a specific regulator of shoot and floral meristem development. We demonstrate that the strong clv3-2 mutant is largely epistatic to clv1 mutants, and that the semi- dominance of clv1 alleles is enhanced by double heterozygosity with clv3 alleles, suggesting that these genes work in the same pathway to control meristem development. We propose that CLV1 and CLV3 are required to promote the differentiation of cells at the shoot and floral meristem.

Self perpetuation of the shoot meristem is essential for the repetitive initiation of shoot structures during plant development. In Arabidopsis shoot meristem maintenance is disrupted by recessive mutations in the WUSCHEL (WUS) gene. The defect is evident at all developmental stages and is restricted to shoot and floral meristems, whereas the root meristem is not affected. wus mutants fail to properly organize a shoot meristem in the embryo. Postembryonically, defective shoot meristems are initiated repetitively but terminate prematurely in aberrant flat structures. In contrast to wild-type shoot meristems, primordia initiation occurs ectopically across mutant apices, including the center, and often new shoot meristems instead of organs are initiated. The cells of wus shoot apices are larger and more vacuolated than wild-type shoot meristem cells. wus floral meristems terminate prematurely in a central stamen. Double mutant studies indicate that the number of organ primordia in the center of wus flowers is limited, irrespective of organ identity and we propose that meristem cells are allocated into floral whorl domains in a sequential manner. WUS activity also appears to be required for the formation of supernumerary organs in the center of agamous, superman or clavata1 flowers, suggesting that the WUS gene acts upstream of the corresponding genes. Our results suggest that the WUS gene is specifically required for central meristem identity of shoot and floral meristems to maintain their structural and functional integrity.

Eukaryotic genes are packaged into dynamic but stable chromatin structures to deal with transcriptional reprogramming and inheritance during development. Chromatin remodeling factors and histone chaperones are epigenetic factors that target nucleosomes and/or histones to establish and maintain proper chromatin structures during critical physiological processes such as DNA replication and transcriptional modulation. Root apical meristems are vital for plant root development. Regarding the well-characterized transcription factors involved in stem cell proliferation and differentiation, there is increasing evidence of the functional implications of epigenetic regulation in root apical meristem development. In this review, we focus on the activities of chromatin remodeling factors and histone chaperones in the root apical meristems of the model plant species Arabidopsis and rice.

Effective somatic cell synchronization in root-tip meristems and improved chromosome spreading were achieved in white campion, wheat, rye, and barley by application of hydroxyurea and amiprophos-methyl or colchicine, combined with a pretreatment of ice water and modified fixative, as well as enzymatic digestion of the meristems. The protocol provides metaphase indices of approximately 50%. The chromosomes and chromosomal DNA were with minimum distortion, providing useful material for chromosome banding studies, in situ DNA–DNA hybridization, microdissection, and microcloning.Key words: Melandrium album, rye, wheat, barley, cell cycle, root meristem, synchronization, metaphase index, chromosome preparation.

In this paper we describe the expression patterns of a family of homeobox genes in maize and their relationship to organogenic domains in the vegetative shoot apical meristem. These genes are related by sequence to KNOTTED1, a gene characterized by dominant neomorphic mutations which perturb specific aspects of maize leaf development. Four members of this gene family are expressed in shoot meristems and the developing stem, but not in determinate lateral organs such as leaves or floral organs. The genes show distinct expression patterns in the vegetative shoot apical meristem that together predict the site of leaf initiation and the basal limit of the vegetative ‘phytomer’ or segmentation unit of the shoot. These genes are also expressed in the inflorescence and floral meristems, where their patterns of expression are more similar, and they are not expressed in root apical meristems. These findings are discussed in relation to other studies of shoot apical meristem organization as well as possible commonality of homeobox gene function in the animal and plant kingdoms.

Plant development is characterized by repeated initiation of meristems, regions of dividing cells that give rise to new organs. During lateral root (LR) formation, new LR meristems are specified to support the outgrowth of LRs along a new axis. The determination of the sequential events required to form this new growth axis has been hampered by redundant activities of key transcription factors. Here, we characterize the effects of three PLETHORA (PLT) transcription factors, PLT3, PLT5, and PLT7, during LR outgrowth. In plt3plt5plt7 triple mutants, the morphology of lateral root primordia (LRP), the auxin response gradient, and the expression of meristem/tissue identity markers are impaired from the “symmetry-breaking” periclinal cell divisions during the transition between stage I and stage II, wherein cells first acquire different identities in the proximodistal and radial axes. Particularly, PLT1, PLT2, and PLT4 genes that are typically expressed later than PLT3, PLT5, and PLT7 during LR outgrowth are not induced in the mutant primordia, rendering “PLT-null” LRP. Reintroduction of any PLT clade member in the mutant primordia completely restores layer identities at stage II and rescues mutant defects in meristem and tissue establishment. Therefore, all PLT genes can activate the formative cell divisions that lead to de novo meristem establishment and tissue patterning associated with a new growth axis.

A number of viruses cause considerable yield loss and quality deterioration in garlic. Root meristems of virus infected plants are known to be free from detectable viruses. This potentiality could be exploited to obtain virus free clones at a high frequency by culturing excised root meristems in vitro. We have developed efficient methods of direct and somatic embryo derived shoot regeneration from root meristems of garlic. The objectives of this work were to detect viruses infecting Bangladeshi and Japanese garlic clones and find an easy and efficient method of eliminating the viruses for the improvement of both yield and quality of garlic. At first, we confirmed the presence of detectable viruses in three Bangladeshi and one Japanese clones. The clones were infected with four different types of viruses: Garlic viruses (GarVs), Onion yellow dwarf virus (OYDV), Leek yellow stripe virus (LYSV), and Garlic common latent virus (GCLV). To eliminate those viruses, as per our previous method, root meristems were cultured on MS medium supplemented with 1.0 µM NAA and 10.0 µM BA. Shoot primordia developed from the cultured explants within 1 month. The regenerated individual shoot buds (2-5 mm) were separated from the mother explants and transferred to growth regulators free medium. RT-PCR confirmed that the viruses present in the mother garlic plants were absent in the shoots found after two-step culture. The regenerated shoots were rooted on growth regulator free medium and transferred to pots. Results indicated that the plants remained free from LYSV. Virus elimination through root meristem culture emerged as an efficient novel technique for the eradication of multiple viruses as confirmed by RT-PCR in this study. This technique has the potential for the production and supply of virus free propagules (plants/bulblets) for the yield and quality improvement of garlic.Progressive Agriculture 28 (2): 55-63, 2017

In both radish and Arabidopsis, lateral root initiation involves a series of rapid divisions in pericycle cells located on the xylem radius of the root. In Arabidopsis, the number of pericycle cells that divide to form a primordium was estimated to be about 11. To determine the stage at which primordia are able to function as root meristems, primordia of different stages were excised and cultured without added hormones. Under these conditions, primordia that consist of 2 cell layers fail to develop while primordia that consist of at least 3–5 cell layers develop as lateral roots. We hypothesize that meristem formation is a two-step process involving an initial period during which a population of rapidly dividing, approximately isodiametric cells that constitutes the primordium is formed, and a subsequent stage during which meristem organization takes place within the primordium.

The growth of a plant's root system depends on the continued activity of the root meristem, and the generation of new meristems when lateral roots are initiated. Plants have developed intricate signalling systems that employ secreted peptides and plasma membrane-localized receptor kinases for short- and long-range communication. Studies on growth of the vascular system, the generation of lateral roots, the control of cell differentiation in the root meristem and the interaction with invading pathogens or symbionts has unravelled a network of peptides and receptor systems with occasionally shared functions. A common theme is the employment of conserved modules, consisting of a short signalling peptide, a receptor-like kinase and a target transcription factor, that control the fate and proliferation of stem cells during root development. This review intends to give an overview of the recent advances in receptor and peptide ligand-mediated signalling involved in root development.

The three-dimensional (3D) arrangement of cells in tissues provides an anatomical basis for analyzing physiological and biochemical aspects of plant and animal cellular development and function. In this study, we established a protocol for tissue clearing and 3D imaging in rice. Our protocol is based on three improvements: clearing with iTOMEI (clearing solution suitable for plants), developing microscopic conditions in which the Z step is optimized for 3D reconstruction, and optimizing cell-wall staining. Our protocol successfully 3D imaged rice shoot apical meristems, florets, and root apical meristems at cellular resolution throughout whole tissues. Using fluorescent reporters of auxin signaling in rice root tips, we also revealed the 3D distribution of auxin signaling events that are activated in the columella, quiescent center, and multiple rows of cells in the stele of the root apical meristem. Examination of cells with higher levels of auxin signaling revealed that only the central row of cells was connected to the quiescent center. Our method provides opportunities to observe the 3D arrangement of cells in rice tissues.

Continuing root growth is crucial for the ongoing survival of a plant and provides the structure basis for the acquisition of valuable resources such as water and nutrients required for growth and development. The root apical meristem, located at the root apex, contains a stem cell niche which is the source of root cell production and patterning. This project focuses on the influence of phytohormone signalling upon the maintenance of the root apical meristem (RAM) in Arabidopsis thaliana. Through a combination of physiological, genetic and molecular approaches, the project has uncovered evidence of the involvement of three plant hormones, auxin, abscisic acid (ABA) and ethylene, in regulating the differentiation of stem cells and their descendants in Arabidopsis root meristems. Key findings include: 1) exogenous application of auxin, ABA or 1-aminocyclopropane-1-carboxylic acid (ACC), which is an ethylene precursor, rescues the root meristem failure of a1f3-1 seedlings; 2) exogenous application of ABA promotes QC quiescence and suppresses stem cell differentiation in wild type Arabidopsis root meristems; 3) inhibition of ethylene biosynthesis and mutations that cause ethylene insensitivity induce differentiation of stem cells in root meristems and.i.on the other hand, exogenous application of ethylene precursor or mutations causes ethylene over-production suppress stem cell differentiation in Arabidopsis root meristems; 4) ABA and ethylene interact antagonistically with auxin in the regulation of both stem cell differentiation in Arabidopsis root meristem, and the expression of the CLE40 gene, which encodes a negative regulator of stem cell differentiation in root meristems; 5) ABA modulates the abundance of the LM6 arabinan cell wall epitope within at the root meristem; 6) ethylene suppresses the stem cell differentiation in Arabidopsis root meristems induced by nitrogen (N) and phosphorus (P) deficiencies. The results of the project demonstrate that stem cell regulation in Arabidopsis root meristems involves complex interactions of plant hormones and environmental signals.

La différentiation cellulaire, qui désigne la transition d’une cellule d’un état souche vers un état mature, est un processus morphogénétique : elle s’accompagne de changements phénotypiques cellulaires issus de modifications de l’expression des gènes. L’activation et la répression de l’expression des gènes résulte de l’activité conjointe de facteurs de transcription et de complexes régulateurs de la chromatine qui instruisent des états chromatiniens locaux via des modifications post-traductionnelles (PTM) des histones nucléosomales. Ces états peuvent faciliter ou empêcher la machinerie de transcription de se lier aux séquences promotrices et régulatrices de gènes. Le Complexe Répressif Polycomb 2 (PRC2) est un complexe régulateur de la chromatine spécialisé dans le dépôt de groupes tri-méthyl sur la lysine 27 de l’histone 3 (H3K27me3). H3K27me3 est en général associée à la répression transcriptionnelle. La perte d’activité de PRC2 désorganise les processus développementaux dans le temps et dans l’espace aux échelles cellulaire et tissulaire. Mon projet doctoral vise à étudier le rôle de PRC2 dans la régulation transcriptionnelle au cours de la différentiation cellulaire lors du développement végétatif de la racine d’Arabidopsis thaliana. En intégrant des données épigénomiques et transcriptomiques en cellule unique, publiées et originales, j'ai disséqué les variations d’expression des gènes ciblés par PRC2 au cours de la différentiation de plusieurs types cellulaires de la racine. Je montre d’abord que la régulation transcriptionnelle par PRC2 est essentiellement type cellulaire-spécifique. De fait, l’activité différentielle des gènes ciblés par PRC2 peut être considérée comme une signature de chaque type cellulaire. De plus, les changements d’activité transcriptionnelle au cours de la différentiation surviennent par vagues spécifiques de chaque type cellulaire et auxquelles contribuent significativement les gènes ciblés par PRC2. La seconde partie de ce projet vise à établir une relation de causalité entre l’activité chromatinienne de PRC2, la régulation transcriptionnelle découlant de l’activité de PRC2 et l’exécution correcte de la différentiation lors de la morphogenèse racinaire. Pour cela, une approche de génétique inverse consistant à implémenter chez A. thaliana un système inductible qui génère, dans des populations de cellules et à un stade développemental choisis, la perte de fonction du gène FIE codant pour une sous-unité essentielle de PRC2. D’après nos premières observations, la perte de FIE post-germination phénocopie des mutants constitutifs de PRC2, suggérant l’implication de PRC2 dans l’orchestration de la différentiation, le maintien des niches de cellules souches et la zonation fonctionnelle des méristèmes racinaires.Notre travail met en lumière l’interaction entre régulation chromatinienne par le dépôt d’une PTM d’histone spécifique, H3K27me3, et la régulation transcriptionnelle au cours de la différentiation cellulaire
Cell differentiation, the process that refers to the transition of stem cells to mature cells, is a morphogenetic process resulting in phenotypic changes at the cellular scale. It relies on a profound remodeling of cells transcriptome. Transcription activation and repression are the result of the intertwined activity of both transcription factors and chromatin-modifying complexes that define local chromatin states by depositing post-translational modifications on histone tails, thereby preventing or facilitating the transcription machinery to bind promoters and other regulatory elements. Polycomb Repressive Complex 2 (PRC2) is a chromatin-modifying complex that catalyses the tri-methylation of lysine 27 of histone 3 (H3K27me3) which deposition is associated with repressive chromatin and transcriptional silencing in eukaryotes. Loss of PRC2 activity deeply impacts developmental processes in plants and metazoans, impairing the orchestration of developmental programs in space and in time, at both cellular and tissular scales.My doctoral project aimed at deciphering the role of PRC2 in regulating transcription during the establishment of cell types during post-embryonic development, using the root of Arabidopsis thaliana as model.By integrating both publicly available and original epigenomic and transcriptomic data at the single cell resolution, I dissected the transcriptional response of PRC2-regulated genes all along the differentiation of several root cell types. These analyses first showed that the transcriptional regulation by PRC2 is for the most part cell type-specific and subsequently that the differential expression of PRC2 target genes is a signature of cell types. Moreover, we found that PRC2-regulated genes are dynamically expressed during cell differentiation and that transcriptional changes occur by waves at key stages in the differentiation of each cell type. The second part of this project seeked to establish a direct causal link between PRC2 activity, the resulting transcriptional regulations and the acquisition of cell identities in A. thaliana primary and lateral roots using a reverse genetics approach. We implemented an inducible gene editing system to knock-out FIE, a gene encoding for a core PRC2 subunit, in a cell type- and developmental stage-specific manner. Using this method, we provide the first evidence of causality between PRC2 activity and its involvement in the homeostasis of root cell differentiation during the post-embryonic development of A. thaliana. Preliminary results showed that FIE knock-out after germination phenocopies classical PRC2 mutants, highlighting the role of PRC2 in both guiding root differentiation and maintaining the indeterminacy and the longitudinal patterning of root meristems to support continuous root growth. Taken together, our results shed new lights into the role of chromatin regulation by PRC2 in the transcriptional control of cell differentiation

Durante esse trabalho de pesquisa verificou-se que a aquisição de competência para conversão de ápices radiculares de Catasetum fimbriatum em gemas caulinares aumentava à medida que as plantas envelheciam. Esse processo esteve relacionado ao estabelecimento do crescimento determinado das raízes e com a parada da atividade e re-organização estrutural do meristema apical radicular (MAR). Este, quando ainda jovem e destituído de competência para a conversão em gemas, apresentava uma organização do tipo fechada, ao passo que em estágios avançados do envelhecimento este padrão transformou-se em um tipo aberto, marcado pela diferenciação e predominância de células parenquimáticas. Tais alterações, aparentemente, ocorreram com a concomitante perda das características e funções do centro de quiescente (CQ). De maneira complementar, constatou-se que a aquisição de competência do MAR para conversão em gemas estava correlacionada a uma série de alterações metabólicas, as quais, supostamente, participaram de uma condição fisiológica favorável a esse processo. Com base no conjunto de dados obtidos, pode-se observar que os teores endógenos de importantes participantes na progressão de divisões celulares, tais como auxinas, citocininas e formas reduzidas de ascorbato e glutationa tenderam a diminuir durante o envelhecimento das raízes. Por outro lado, durante esse mesmo período, o conteúdo de alguns hormônios envolvidos na sinalização de condições de estresse ou diferenciação celular, tais como etileno, ácido abscísico e giberelinas tenderam a aumentar. As concentrações relativas de importantes sinalizadores secundários, tais como óxido nítrico e cálcio citossólico também apresentaram aumento conspícuo na região do MAR durante o envelhecimento. Agregando elementos a estas constatações, verificou-se que o transporte polar de auxina seria um importante sinal posicional para a manutenção das características e função do MAR, uma vez que o seu bloqueio em plantas jovens foi suficiente para causar a aquisição da competência do MAR, no entanto, o processo de conversão não era consolidado enquanto os ápices radiculares permaneceram ligados às plantas. A aplicação de etileno em plantas jovens, por sua vez, desencadeou efeitos similares; no entanto, além de induzir a competência, esse hormônio também proporcionou a conversão dos MARs em gemas via aumento nos teores endógenos de citocininas. O tratamento de ápices radiculares jovens com diferentes tipos de citocininas revelaram que citocininas do tipo isopenteniladenina (iP e iPR) mostraram-se mais de perto relacionadas à retenção de características radiculares, ao passo que as do tipo zeatina (Z e ZR) apresentou maior influência e presença em condições em que as características radiculares foram perdidas. Por outro lado, a aplicação de substâncias moduladoras do balanço redox em ápices radiculares jovens mostrou que o estresse oxidativo proporcionou a aquisição de competência do MAR para conversão em gemas. Essa mesma tendência foi observada com a aplicação de concentrações relativamente elevadas de substâncias indutoras da elevação dos teores de óxido nítrico e cálcio citossólico nos tecidos. Os ápices radiculares com competência parcialmente estabelecida, analisados logo nas primeiras horas após o isolamento, revelaram que sua separação da planta-mãe acelerava as mudanças morfológicas que naturalmente ocorrem no MAR em estágios avançados do envelhecimento. Durante esse mesmo período, verificou-se uma queda rápida nos teores endógenos de citocininas (principalmente do tipo iP), proporcionando a predominância de citocininas do tipo Z durante a maior parte do primeiro dia de isolamento dos explantes, a qual coincidiu com a mudança no padrão de organização do MAR do tipo intermediário-aberto para o totalmente aberto. O avanço das modificações no ápice radicular após esse período desencadeou o estabelecimento do meristema caulinar, cujo evento esteve relacionado a uma tendência de aumento nos teores de citocininas e de ascorbato após o primeiro dia de isolamento. Dessa forma, os estágios mais avançados do envelhecimento radicular, bem como a separação de ápices radiculares com competência parcialmente estabelecida, parecem desencadear e aumentar a competência do MAR para conversão por meio de modificações morfológicas e fisiológicas muito similares nos ápices radiculares. Essas alterações envolveram a perda das características radiculares, a qual parece depender de alterações no controle exercido pelo CQ sobre o desenvolvimento das demais células no MAR. Esses eventos na região do CQ se revelaram condição sine qua non para a complementação da aquisição de competência do MAR, sendo esta dependente da intensidade das perturbações sobre o controle da organização do MAR. Dessa forma, a conversão do meristema apical radicular de C. fimbriatum em gemas caulinares parece decorrer da formação de um novo grupo de células na antiga região do CQ do MAR alterado. Essas células pareceram competentes para responder a diferentes estímulos que as direcionariam a uma nova rota do desenvolvimento que, nesse caso, seria o estabelecimento de um meristema caulinar com conseqüente desenvolvimento de uma gema vegetativa.
During this research work, it was noticed that competence acquisition for the conversion of Catasetum fimbriatum root tips into buds was related to the plant ageing. This process seems to be coupled with the establishment of the determinate root growth and with the cessation in the activity and structural re-organization of the root apical meristem (RAM). Young and non-competent root tips showed a closed RAM architecture, and the ageing process stimulated the establishment of an open organization in the RAM, as indicated by a higher level of differentiation and a predominance of parenchymatic cells in the old root apices. These alterations were concomitant with the modifications on the characteristics and functions of quiescent center (QC). In agreement with these observations, the competence acquisition to the conversion of the MAR into buds was linked to a series of metabolic alterations, which probably play a role in this process. Based on the data obtained, it was observed that the endogenous levels of important components of the cell division progression, such as auxins, cytokinins and the reduced forms of ascorbate and glutathione showed a tendency of decrease during the root ageing. On the other hand, during this same period, the content of some hormones involved in signalling events of stress conditions or cellular differentiation, such as ethylene, abscisic acid and gibberellins exhibited a pattern of increase. The relative concentrations of important second messengers, such as nitric oxide and cytosolic calcium also displayed a marked increased in the RAM region during the ageing. Additionally, it was noticed that the auxin polar transport represents an important positional signal for the maintenance of the RAM characteristics and functions, once treatments that blocked the transport of this hormone promoted the MAR competence acquisition even in young plants, although, the conversion process did not complete while the root tips were maintained attached to the plants. The treatment of young plants with ethylene, on the contrary, caused similar effects; however, besides inducing the competence, this hormone also promoted the RAM conversion into buds via the elevation in the endogenous levels of cytokinins. The treatment of young root tips with different types of cytokinins indicated that iP-type cytokinins (iP and iPR) were more closely associated to the preservation of the root characteristics, while the Z-type cytokinins (Z and ZR) showed a higher importance when the root characteristics were lost. Furthermore, the treatment of young root apices with compounds that cause alterations in the cellular redox status indicated that the oxidative stress stimulated the competence acquisition for the RAM conversion into buds. This same tendency was observed with the application of relatively high concentrations of compounds that induce elevations in the levels of nitric oxide and cytosolic calcium in the tissues. The analyses carried out during the first hours after the isolation of partially competent root apices indicated that the detachment of the root tips from the original plants accelerated the morphological modifications that naturally occur at advanced stages of ageing. During this same period, it was observed a rapid decrease in the endogenous levels of cytokinins (specially of the iP-type), leading to a predominance of the Z-type cytokinins during the first day after the isolation of the explants, which coincided with the alteration of the RAM architecture from the intermediate-open type to the completely open type. After the first day of isolation, the progress in the root apices modifications resulted in the establishment of the shoot meristem, which was accompanied by an elevation in the endogenous levels of cytokinins and ascorbate. Therefore, advanced stages of root ageing, as well the isolation of the partially competent root apices, seem to increase the competence for the RAM conversion into buds via similar morphological and physiological changes in the root apices. These alterations involved the loss of the root characteristics, which possibly resulted from modifications in the control of the QC on the development of the other cells in the RAM. These events in the QC represent a sine qua non condition for the completion of the MAR competence acquisition, which is affected by the intensity of the perturbations on the control of the RAM organization. Therefore, the conversion of root apical meristem of C. fimbriatum into buds probably results from the formation of a new group of cells in the region of the QC of the altered RAM. These cells seem to be competent to respond to different stimulus that would directionate them to a new developmental route that, in this case, consists in the establishment of a shoot meristem.

Setaria viridis é uma espécie de gramínea muito relevante em estudos de desenvolvimento vegetal, por determinadas características que a fazem uma excelente proposta de organismo modelo para plantas monocotiledôneas de metabolismo C4. Para que a espécie seja utilizada amplamente em estudos que visem compreender o funcionamento e desenvolvimento vegetal, bem como os mecanismos moleculares que o modulam, é essencial que aspectos de seu desenvolvimento sejam desvendados. Nesse contexto, a caracterização do meristema radicular e das estruturas presentes nos estágios iniciais da germinação é importante para se compreender como ocorre, em gramíneas, o surgimento de tipos de raízes diferentes, além de correlacionar quais fatores ambientais e endógenos estão envolvidos na escolha de diferentes arquiteturas de sistemas radiculares em gramíneas. Além disso,a caracterização do desenvolvimento de estruturas florais em Setaria viridis é importante, uma vez que traz informações que podem contribuir com um aumento na eficiência de metodologias de transformação genética para a espécie, via \"spike dip\". O presente trabalho caracterizou morfoanatomicamente o desenvolvimento radicular desde os estágios iniciais da germinação e o desenvolvimento floral em Setaria viridis. Além disso, buscou estabelecer relação entre o surgimento de diferentes tipos de raízes (primárias e as adventícias) com diferentes condições de luminosidade em cultivos in vitro
Setaria viridis is a very important grass in studies of plant development, due to some characteristics that make it an excellent proposed model organism for monocotyledonous plants with C4 metabolism. To be widely used in studies that aim to understand plant functioning and development, as well as the molecular mechanisms that modulate it, it is essential that aspects of its development be unraveled. In this context, the characterization of the root meristem and structures present in the early stages of germination is also important to understand how the emergence of different root types occurs in grasses. Moreover, to correlate the environmental and endogenous factors involved in the choice of different architectures of root systems in grasses. Furthermore, the characterization of the development of floral structures in Setaria viridis is important, since it brings information that can contribute to the efficiency of methodologies of genetic transformation for the species. The present work characterized morphologically the root development during the initial stages of germination and the floral development of Setaria. In addition, it sought to establish a relationship between the emergences of different types of roots (primary and adventitious) with different light conditions in in vitro cultures

Dans l’optique d’une seconde Révolution Verte, visant, à la différence de la première, à accroître les rendements des cultures dans un contexte de faible fertilité, les stratégies mises en place par les plantes pour une assimilation optimale des nutriments du sol se trouvent au cœur du problème. Afin de le résoudre et d’identifier les variétés idéales parmi la diversité génétique des plantes cultivées, les systèmes racinaires, leur développement et leur architecture, sont appelés à jouer le premier rôle. La variabilité au sein des racines latérales semble s’avérer une caractéristique cruciale pour l’optimisation de l’exploration du sol et de l’acquisition de ses ressources mobiles et immobiles, mais ce phénomène est encore mal appréhendé.Le travail présenté dans cette thèse se concentre sur les racines latérales du maïs (Zea mays L.) dans un effort pour révéler les processus à l’origine des variations intrinsèques dans le développement racinaire. Il s’appuie en particulier sur le phénotypage des racines latérales à une échelle sans précédent, suivant la croissance journalière de milliers d’entre elles à haute résolution spatiale, pour caractériser précisément les variations spatio-temporelles entre et au sein des individus racinaires. Les profils individuels de vitesse de croissance ont été analysés à l’aide d’un modèle statistique qui a identifié trois principales tendances temporelles dans les vitesses de croissance menant à la définition de trois classes de racines latérales avec une vitesse et durée de croissance distinctes. Des différences de diamètre à l’émergence de ces racines (dont l’origine remonte au stade du primordium) conditionnent probablement la tendance ultérieur de croissance mais ne suffisent pas à déterminer le destin de la racine. Finalement, ces classes racinaires sont distribuées aléatoirement le long de la racine primaire, ce qui suggère qu’aucune stimulation ou inhibition locale n’existe entre racines voisines.Pour expliquer l’origine des variations observées dans la croissance, ce travail a été complété par une caractérisation multi-échelle de groupes de racines latérales présentant une croissance distincte, à un niveau cellulaire, anatomique et moléculaire. Un effort particulier a été dirigé à l’analyse des profils de longueur de cellules dans des apex racinaires pour lequel nous avons introduit un modèle de segmentation pour identifier des zones développementales. Grâce à cette méthode, une forte modulation dans la longueur des zones de division et d’élongation a été mise en évidence, en lien avec les variations de la croissance des racines latérales. Le rôle régulateur de l’auxine sur l'équilibre entre les processus de prolifération et d’élongation cellulaire a été montré avec l’utilisation de lignées mutantes. En fin de compte, les variations de la croissance entre racines latérales sont remontées jusqu’à l’allocation d’assimilats carbonés et la capacité de transport de la racine, ce qui suggère l’existence d’un mécanisme de rétroaction qui pourrait jouer un rôle déterminant dans la mise en place de tendances contrastées dans la croissance des racines latérales
In the perspective of a second Green Revolution, aiming, unlike the first one, to enhance yields of crops in a low fertility context, the strategies used by plants for an optimal uptake of soil nutrients are at the core of the problem. To solve it and identify ideal breeds among the genetic diversity of crops, plant root systems, their development and their architecture, are called upon to play the leading role. The variability among secondary roots appears as a crucial feature for the optimality of soil exploration and acquisition of mobile and immobile resources, but this phenomenon remains poorly understood. The work presented in this thesis focuses on the lateral roots of maize (Zea mays L.) and attempts to unravel the processes at the origin of intrinsic variations in lateral root development. It relies notably on the phenotyping of individual lateral roots at an unprecedented scale, tracking the daily growth of thousands of them at a high spatial resolution, in order to characterize precisely the spatio-temporal variations existing both between and within root individuals. Individual growth rate profiles were analyzed with a statistical model that identified three main temporal trends in growth rates leading to the definition of three lateral root classes with contrasted growth rates and growth duration. Differences in lateral root diameter at root emergence (originating at the primordium stage) were likely to condition the followed growth trend but did not seem enough to entirely determine lateral root fate. Lastly, these lateral root classes were randomly distributed along the primary root, suggesting that there is no local inhibition or stimulation between neighbouring lateral roots. In order to explain the origin of the observed differences in growth behaviour, we complemented our study with a multi-scale characterization of groups of lateral roots with contrasted growth at a cellular, anatomical and molecular level. A particular focus is set on the analysis of cell length profiles in lateral root apices for which we introduced a segmentation model to identify developmental zones. Using this method, we evidenced strong modulations in the length of the division and elongation zones that could be closely related to variations in lateral root growth. The regulatory role of auxin on the balance between cellular proliferation and elongation processes is demonstrated through the analysis of mutant lines. Ultimately, variations in lateral root growth are traced back to the allocation of carbon assimilates and the transport capacity of the root, suggesting that a feedback control loop mechanism could play a determinant role in the setting out of contrasted lateral root growth trends

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Conselho Nacional de Desenvolvimento Científico e Tecnológico
A organogênese de brotos a partir de raízes (rootsuckers) permite a propagação vegetativa daArabidopsis lyrata, o parente mais próximo daArabidopsis thaliana. Utilizando um sistema in vitro, o presente estudo objetivou compreender melhor a propagação vegetativa nessa espécie modelo A. lyrata, no que se refere ao desenvolvimento morfológico de suckers, à capacidade de propagação vegetativa em diferentes condições de crescimento in vitro e à identificação de genes potencialmente envolvidos na formação do meristema apical dos brotos.O surgimentodos suckers ocorreu após 30 dias, mais freqüentemente na região axilar das raízes laterais. Os cortes transversais das raízes mostraram uma estrutura primária típica diarca e após cerca de 25 dias, pode-se observar o crescimento secundário da raiz, como indicado pela formação do câmbio. Conclui-se que a emergência do sucker assemelha-se à iniciação das raízes laterais a partir do periciclo, tecido que dá origem ao câmbio vascular durante o crescimento secundário. Em relação às condições de crescimento in vitro, a força total no meio MS induziu o maior número de suckers por planta, seguido por alta concentração de sacarose (3%).Exposição à luz e privação de sacarose não são estritamente necessários para a formação de suckers. Nossos dados também revelaram que a auxina promove a formação dos brotos. Máximas de auxina vascular são necessários para desencadear a iniciação da raiz lateral, sugerindo que a formação de suckers promovida por auxina ocorre provavelmente por mecanismos semelhantes. A avaliação de diferentes genes relacionados a meristema apical, demonstram que o gene STM pode ser um marcador para distinguir as células responsáveis pela formação de suckers. Arabidopsis lyrata provou ser um excelente modelo para estudos de organogênese em raíz e posteriores estudos usando esse sistema de reproduçãopara detectar marcadores epigenéticos através das várias gerações de propagação clonal.
Shoot organogenesis from roots (root suckers) allows vegetative propagation of Arabidopsis lyrata, the closest relative of Arabidopsis thaliana, in addition to sexual propagation and is an important trait associated with the root system. Using an in vitro system, we aimed to better understand the vegetative propagation in the model species A. lyrata, in what regards the morphological development of root suckers, the ability of vegetative propagation in different in vitro growth conditions, and identifying genes potentially involved in the formation of the new shoot apical meristem.Root sucker appearanceoccurred after30 days,most frequently in the axils of lateral roots. Root cross-sections showed a typical diarch primary structure and after 25 days, secondary root growth could be observed, as indicated by formation of the cambium. According to our data,root sucker emergence resembles the initiation of lateral roots from the pericycle, the tissue that gives rise to the vascular cambium during secondary growth. Regarding the in vitro growth conditions, full strength of MS induced the highest number of root suckers per plant, followed 3% of sucrose. However, light exposure and sucrose deprivation are not strictly required for sucker formation. Our data also revealed that auxin promotes root suckering. Vascular auxin response maxima are required to trigger lateral root initiation, suggesting that auxin- promoted sucker formation likely occurs by similar mechanisms. The evaluation of different shoot apical meristem related genes, suggests that the STM gene can be a potential marker to identify cells responsible in driving sucker formation. Arabidopsis lyrata proved to be an excellent model for further studies using root suckers, for example to study epigenetic marks throughout generations of clonal propagation.

L'auxine régule la croissance et le développement des plantes grâce aux facteurs de transcription de la famille des "AUXIN RESPONSE FACTOR" (ARF). Chez Arabidopsis thaliana en particulier, ARF5, 6, 7, 8 et 19 activent l'expression de gènes cibles en réponse à l'auxine. Ces cinq ARF activateurs contrôlent de façon plus ou moins redondante des processus divers au cours du développement de la plante, notamment la régulation des croissances au niveau des méristèmes racinaires et aériens ainsi que la formation des racines latérales ou des méristèmes axillaires.Chacun de ces cinq ARF activateurs présente des patrons d'expression uniques dans chacun des tissus racinaires et aériens, en association avec leurs fonctions particulières. Il est probable que cette expression tissu-spécifique trouve son origine dans un contrôle différencié de leur transcription. Dans cette étude, des régulateurs amonts de la transcription de ARF5, 6, 7, 8 et 19 ont été identifiés par une méthode haut-débit de crible simple hybride en levure (Y1H). Une procédure d'expression transitoire en protoplastes a permis de confirmer que l'expression de chaque ARF activateur est contrôlée par des régulateurs spécifiques, dont la majorité se comportent comme des répresseurs de la transcription des ARF in planta. Parmi les régulateurs identifiés, les facteurs de transcription ont été étudiés grâce à des mutants pour préciser les interactions in planta. Ces mutants montrent des phénotypes développementaux typiques de perturbations de l'auxine dans les racines et les tiges : altérations des cinétiques de croissance, de l'émergence des organes latéraux ou de réponses à l'auxine et modification de l'expression des ARF activateurs.Par ailleurs, ce travail aborde également les dialogues entre les voies de signalisation de l'auxine et des cytokinines, et en particulier le rôle de ces interactions dans le développement des racines et des tiges. Une des interactions identifiées dans le crible Y1H est la répression de ARF7 par CRF10, un gène membre de la famille des "Cytokinin Response Factors". Nous avons mis en évidence l'importance de cette interaction pour le maintien de l'architecture du méristème apical racinaire, pour la sénescence des feuille et pour la réponse phototropique à la lumière bleue dans les hypocotyles
Auxin regulates plant growth and development through the transcription factors of the AUXIN RESPONSE FACTOR (ARF) gene family. Most notably in Arabidopsis thaliana ARF5, 6, 7, 8 and 19 activate expression of target genes in response to auxin. These five ARF activators control both variable and overlapping processes during plant development including regulation of growth at the root and the shoot apical meristems, lateral root and axillary shoot formation. Each of the five ARF activators shows unique tissue-specific expression patterns in the root and the shoot associated with their distinct functions. This tissue-specific expression is likely derived from the differences in the control of ARF activator transcription. In this study the upstream regulators of ARF5, 6, 7, 8 and 19 transcription were identified. This was achieved by utilizing a high-throughput yeast one-hybrid (Y1H) method. The transient protoplast assay revealed that each ARF activator is controlled by specific transcriptional regulators and that the majority of these regulators are repressors of ARF transcription in planta. Mutants of the regulatory transcription factors were utilized to additionally investigate the interactions in planta. These mutants display auxin-related developmental phenotypes in the root and the shoot including alternations in growth kinetics, emergence of lateral organs, responses to auxin and altered expression of ARF activators. Furthermore, this study additionally focuses on cross-talk between the auxin and cytokinin signaling pathways and its role in root and shoot development. One of the interactions identified in the Y1H screen is a repression of ARF7 by CRF10, a member of the Cytokinin Response Factors gene family. The importance of this interaction in maintaining architecture of the root apical meristem, in leaf senescence and in the phototropic response to blue light in hypocotyls is studied

La paroi des cellules végétales subit des modifications afin de s'assouplir ou de se rigidifier selon les besoins de la plante. Cette paroi est une structure complexe, composée de cellulose, d'hémicellulose et de pectines. Les modifications subies par les pectines au cours de l'élongation cellulaire sont encore assez peu caractérisées. Dans ce contexte, le but de ce projet est d'étudier le rôle de deux polygalacturonases (PG) dans le développement racinaire de la plante modèle A. thaliana. Les PG sont des enzymes de dégradation des homogalacturonanes (HG), le composant pectique majoritaire de la paroi primaire. Notrehypothèse est que les PG dégradent partiellement les HG des parois longitudinales des cellules racinaires en élongation. Cette dégradation engendrerait un assouplissement pariétal localisé, permettant la croissance anisotropique des cellules. Nos résultats indiquent que les gènes des deux PG étudiées, nommés PG ROOT APICAL MERISTEM (PG RAM) et PG ROOT (PG R) sontexprimés de façon complémentaire dans la racine, l'un dans le méristème racinaire (PG RAM), et l'autre dans la zone d'élongation et de différenciation (PG R). De plus, la sur-expression de la protéine PG R entraine une augmentation de l’élongation des hypocotyles étiolés, ainsi qu'une augmentation de la densité de racines latérales par rapport au sauvage, démontrant son rôle dans le développement racinaire et dans l'allongement cellulaire. Enfin, nous avons démontré que l'expression des gènes de ces PG était contrôlée de façon différentielle par les facteurs de transcription de la famille PLETHORA (PLT)
Plant cell wall structure is modified to control its stiffness or flexibility according to plant’s requirements. The cell wall is a complex structure, composed of cellulose, hemicelluloses and pectins. Pectin modifications during cellular elongation are not very well characterized. In this context, the aim of this project is to study the roles of two polygalacturonases (PG) in the root development on the model plant A. thaliana. PG are homogalacturonans (HG) degradation enzymes, HG being the major pectic component of the primary cell wall. This degradation would lead to a local parietal relaxation, allowing anisotropic growth of the cells. Our results show that the two studied PG, named PG ROOT APICAL MERISTEM (PG RAM) and PG ROOT (PG R), are expressed in complementary areas of the root, either in the root apical meristem (PG RAM) or in the elongated and differenciated root tissues (PG R). Furthermore, the over-expression of PG R results in longer etiolated hypocotyls and increases root density when compared to wild-type, demonstrating its function in root development and in cell elongation. Finally, we demonstrated that expression of these two PG genes is under the control of PLETHORA (PLT) family transcription factors, by differentially ways

Abstract Root development and growth is similar to shoot growth in that extension growth is initiated by an apical meristem and girth growth of mature roots is carried out by the vascular cambium. However, the initiation of lateral roots is entirely different than the initiation of lateral leaves or shoot meristems. This chapter deals with understanding the root sink in fruit trees by studying root growth, including the initiation of lateral roots, root classification according to size and function, factors affecting their growth, and rootstocks.

The complex use of indirect methods (counting chloroplasts in stomatal cells, as well as direct counting of chromosomes in preparations of root meristems) in combination with DNA methods makes it possible to identify ploidy of plants obtained from white cabbage anthers and to rank them on haploids / doubled haploids and diploid ones already on test tube level.

The impact analysis of low doses of ionizing radiation on the breathers in natural populations is an important part of radiobiological studies of non-human biota. The main aim of our studies was to investigate some cytogenetic, morphological and reproductive rates of the common reed (Phragmites australis (Cav.) Trin. ex. Steud.) from different water bodies within the Chernobyl accident exclusion zone. The absorbed dose rate for littoral emergent plants in sampling water bodies was varied from 1.3E−02 to 1.6E−01 Gy/h. The rate and main types of chromosome aberrations in roots meristems, morphological damages in seed germs, as well as rates of germinating ability and power were analyzed. There were registered rather low rate of germinating ability (14–48%) and germinating power (<1) of seeds from all sampling water bodies with high levels of radioactive contamination in comparison to control ones. Against the general suppressed background the effect of relative stimulation of more affected seeds was observed. With increase of absorbed dose in range of 1.3E−02–1.6E−01 Gy/h the number of germinated seeds was increased. At the same time the number of morphological damages of seeds was increased as well. There was determined the positive correlation between absorbed dose rate and chromosome aberration rate in roots of the common reed from sampling water bodies. The highest rate of chromosome aberrations (up to 17%) were registered in plants with high level of morphological deviations in seeds germs. The data obtained from the complex analysis of natural populations of the common reed from the radioactive contaminated water bodies testify about rather high level of genetic efficiency of low doses of long-term exposure.

The shoot apical meristem establishes plant architecture by continuously producing new lateral organs such as leaves, axillary meristems and flowers throughout the plant life cycle. This unique capacity is achieved by a group of self-renewing pluripotent stem cells that give rise to founder cells, which can differentiate into multiple cell and tissue types in response to environmental and developmental cues. Cell fate specification at the shoot apical meristem is programmed primarily by transcription factors acting in a complex gene regulatory network. In this project we proposed to provide significant understanding of meristem maintenance and cell fate specification by studying four transcription factors acting at the meristem. Our original aim was to identify the direct target genes of WUS, STM, KNAT6 and CNA transcription factor in a genome wide scale and the manner by which they regulate their targets. Our goal was to integrate this data into a regulatory model of cell fate specification in the SAM and to identify key genes within the model for further study. We have generated transgenic plants carrying the four TF with two different tags and preformed chromatin Immunoprecipitation (ChIP) assay to identify the TF direct target genes. Due to unforeseen obstacles we have been delayed in achieving this aim but hope to accomplish it soon. Using the GR inducible system, genetic approach and transcriptome analysis [mRNA-seq] we provided a new look at meristem activity and its regulation of morphogenesis and phyllotaxy and propose a coherent framework for the role of many factors acting in meristem development and maintenance. We provided evidence for 3 different mechanisms for the regulation of WUS expression, DNA methylation, a second receptor pathway - the ERECTA receptor and the CNA TF that negatively regulates WUS expression in its own domain, the Organizing Center. We found that once the WUS expression level surpasses a certain threshold it alters cell identity at the periphery of the inflorescence meristem from floral meristem to carpel fate [FM]. When WUS expression highly elevated in the FM, the meristem turn into indeterminate. We showed that WUS activate cytokinine, inhibit auxin response and represses the genes required for root identity fate and that gradual increase in WUCHEL activity leads to gradual meristem enlargement that affect phyllotaxis. We also propose a model in which the direction of WUS domain expansion laterally or upward affects meristem structure differently. We preformed mRNA-seq on meristems with different size and structure followed by k-means clustering and identified groups of genes that are expressed in specific domains at the meristem. We will integrate this data with the ChIP-seq of the 4 TF to add another layer to the genetic network regulating meristem activity.

Genetic and biochemical analysis of glucosinolate breakdown: The effects of indole-3-carbinol on plant physiology and development Glucosinolates are a class of defense-related secondary metabolites found in all crucifers, including important oilseed and vegetable crops in the Brassica genus and the well-studied model plant Arabidopsis thaliana. Upon tissue damage, such as that provided by insect feeding, glucosinolates are subjected to catalysis and spontaneous degradation to form a variety of breakdown products. These breakdown products typically have a deterrent effect on generalist herbivores. Glucosinolate breakdown products also contribute to the anti-carcinogenic effects of eating cabbage, broccoli and related cruciferous vegetables. Indole-3-carbinol, a breakdown product of indol-3-ylmethylglucosinolate, forms conjugates with several other plant metabolites. Although some indole-3-carbinol conjugates have known functions in defense against herbivores and pathogens, most play as yet unidentified roles in plant metabolism, and possibly also plant development. At the outset, our proposal had three main hypotheses: (1) There is a specific detoxification pathway for indole-3-carbinol; (2) Metabolites derived from indole-3-carbinol are phloem-mobile and serve as signaling molecules; and (3) Indole-3-carbinol affects plant cell cycle and cell-differentiation pathways. The experiments were designed to enable us to elucidate how indole-3-carbinol and related metabolites affect plants and their interactions with herbivorous insects. We discovered that indole-3- carbinol rapidly and reversibly inhibits root elongation in a dose-dependent manner, and that this inhibition is accompanied by a loss of auxin activity in the root meristem. A direct interaction between indole-3-carbinol and the auxin perception machinery was suggested, as application of indole-3-carbinol rescued auxin-induced root phenotypes. In vitro and yeast-based protein interaction studies showed that indole-3-carbinol perturbs the auxin-dependent interaction of TIR1 with Aux/IAA proteins, supporting the notion that indole-3-carbinol acts as an auxin antagonist. Furthermore, transcript profiling experiments revealed the influence of indole-3-carbinol on auxin signaling in root tips, and indole-3-carbinol also affected auxin transporters. Brief treatment with indole-3-carbinol led to a reduction in the amount of PIN1 and to mislocalization of PIN2. The results indicate that chemicals induced by herbivory, such as indole-3-carbinol, function not only to repel herbivores, but also as signaling molecules that directly compete with auxin to fine tune plant growth and development, which implies transport of indole-3- carbinol that we are as yet unsuccessful in detecting. Our results indicate that plant defensive metabolites also have secondary functions in regulating aspects of plant metabolism, thereby providing diversity in defense-related plant signaling pathways. Such diversity of of signaling by defensive metabolites would be beneficial for the plant, as herbivores and pathogens would be less likely to mount effective countermeasures. We propose that growth arrest can be mediated directly by the herbivory-induced chemicals, in our case, indole-3-carbinol. Thus, glucosinolate breakdown to I3C following herbivory would have two outcomes: (1) Indole-3-carbinaol would inhibit the herbivore, while (2) at the same time inducing growth arrest within the plant. Thus, our results indicate that I3C is a defensive phytohormone that modulates auxin signaling, leading to growth arrest.