Submitted by Rafaela Prosdocini Parmeggiani (rafaela.prosdocini@gmail.com) on 2016-02-16T16:01:51Z No. of bitstreams: 1 Atividade cambial em ramos de Kielmeyera grandiflora (Wawra) Saddi (Calophyllaceae) ocorrentes no cerrado paulista.pdf: 2086514 bytes, checksum: d4efa2e5aa9b442218c364e59bc53a9a (MD5) / Approved for entry into archive by Ana Paula Grisoto (grisotoana@reitoria.unesp.br) on 2016-02-17T15:53:14Z (GMT) No. of bitstreams: 1 parmeggiani_rp_me_bot.pdf: 2086514 bytes, checksum: d4efa2e5aa9b442218c364e59bc53a9a (MD5) / Made available in DSpace on 2016-02-17T15:53:14Z (GMT). No. of bitstreams: 1 parmeggiani_rp_me_bot.pdf: 2086514 bytes, checksum: d4efa2e5aa9b442218c364e59bc53a9a (MD5) Previous issue date: 2015-12-16 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / No cerrado, as estações são bem demarcadas pela sazonalidade pluvial, portanto, espera-se que haja uma sazonalidade na atividade do câmbio vascular. Kielmeyera grandiflora é considerada uma espécie modelo para estudos de atividade cambial em ramos, pois possui módulos de crescimento bem demarcados, formados por cicatrizes da gema terminal. Nosso objetivo foi de entender qual a relação entre a atividade cambial de Kielmeyera grandiflora (Wawra) Saddi (Calophyllaceae), os fatores ambientais e a fenologia e responder se a atividade cambial varia entre três módulos de crescimentos consecutivos. A área de estudo é um fragmento de cerrado em regeneração no município de Botucatu, estado de São Paulo, Brasil (S 22º53’11.0’’, W 48º29’17.3’’). Foram amostrados mensalmente os três últimos módulos de crescimento dos ramos de três indivíduos durante o período de fevereiro de 2012 a fevereiro de 2013. No mesmo período, a fenologia (brotação, folhas novas, folhas adultas, folhas senescentes e queda foliar) foram avaliadas. O material foi fixado em CRAF III para conservação do conteúdo celular. As amostras foram desidratadas em série alcoólica e incluídas em resina metacrilato para posterior corte em micrótomo rotativo. As análises foram feitas em fotomicroscópio de luz. Utilizamos Modelo Generalizado Linear Misto (GLMM) com erro Binomial para a variável resposta presença/ausência da atividade cambial e erro de Poisson para a variável resposta de contagem de paredes recém formadas. O início da atividade cambial, a brotação e o desenvolvimento de folhas novas ocorreram durante o mês de agosto no período seco. O período de atividade cambial sobrepôs-se à estação chuvosa, temperaturas elevadas e comprimento do dia entre 12 e 13,6 h. O câmbio vascular entrou em dormência em maio, mês que ainda chovia, contudo a temperatura e o comprimento do dia passavam por quedas acentuadas. A variação da atividade cambial entre os módulos de crescimento foi representativa somente no período de reativação cambial, mostrando que o módulo 1, mais próximo às fontes de auxina, apesar de ter iniciado atividade junto ao módulo 3, produziu em todo o período analisado, maior quantidade de células novas, possivelmente devido à ação de hormônios produzidos nos tecidos jovens e em desenvolvimento. / In the cerrado, the seasons are well marked by rain seasonality; therefore, there is a seasonality in the vascular cambium activity. Kielmeyera grandiflora is considered a model for branch cambial activity studies because it has well marked growth modules formed by terminal bud scars. Our goal was to understand what the relationship between cambial activity of Kielmeyera grandiflora (Wawra) Saddi (Calophyllaceae), environmental factors and the phenology and answear wheter cambial activity varies between three consecutive sections of internodal growth. The study area is a cerrado fragment in regeneration in the city of Botucatu, São Paulo State, Brazil (22° 53 ' S 11.0 '', W 48° 29 ' 17.3 ''). Were sampled on a monthly basis the last three modules of growth of branches of three individuals during the period between February 2012 to February 2013. The material was fixed in CRAF III for preservation of cell content. The samples were dehydrated in alcohol series and included in historesin for further cut in rotary microtome. Analyses were made on light microscope. We used Generalized Linear Mixed Model (GLMM) with Binomial error for the variable reply presence/absence of cambial activity and Poisson error for the variable count response of newly formed walls. The beginning of cambial activity, the sprouting and the development of new leaves occurred during the month of August in the dry period. The period of cambial activity overlapped the rainy season, high temperatures and day length between 12 and 13, 6 h. The vascular cambium dormancy was observed in May, a raining month; however, temperature and day length was decreasing. The variation of cambial activity between the internodal growth sections was representative just in reactivation period, showing that the younger internodal section, closest to the sources of auxin, has produced in the whole study period a larger number of new cells that the older, possibly due to the action of hormones produced in young and development tissues.

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Le genre Larix intéresse les reboiseurs en raison de sa croissance rapide et de son bois apprécié. En particulier, le mélèze est une alternative intéressante au douglas pour les reboisements à basse et moyenne altitude. L'adaptation à l'environnement est une question clé pour les améliorateurs d'arbres forestiers. Cette question est particulièrement importante dans le cadre du changement climatique, où une diminution de la disponibilité en eau durant la saison de végétation est prédite. Trois espèces principales de mélèzes sont utilisées en reboisement en France : le mélèze d'Europe, le mélèze du Japon et leur hybride. Les bonnes performances du mélèze hybride en plantation témoignent de son grand potentiel. La structure cellulaire des cernes annuels reflète la réponse du cambium (division cellulaire, allongement et épaississement des parois) aux facteurs environnementaux (climat, fertilité, compétition…), ainsi que des stades physiologiques et l'effet de la génétique. La densité du bois peut être interprétée comme le produit de l'activité cambiale, et sa variation comme la réponse de l'arbre à son environnement. Toutefois, afin de bien avons combiné de façon originale microdensitométrie et blessures du cambium comprendre cette réponse et les variations du processus de xylogenèse lui-même, il est important de repérer les évènements qui se succèdent lors de la production des cellules de bois. Plusieurs approches permettent de dater la formation du bois. Nous (pinning method) afin d'étudier la dynamique de la fabrication du bois. Nous avons analysé les données obtenues du point de vue de l'améliorateur d'arbres forestiers. Cette nouvelle approche nous a permis d‘aller au-delà le caractère statique du profil microdensitométrique (où la densité varie en fonction de la distance) afin de le convertir en profil dynamique (où la densité varie en fonction du temps). La variation de la phénologie de l'activité cambiale peut être vue comme un mécanisme ajustant bois peuvent être interprétées comme des caractères d'adaptation. L'intérêt pour les les le fonctionnement des arbres à leur environnement. Si cet ajustement améliore leur valeur adaptative (fitness), alors les variations correspondantes de l'anatomie du entre phénologie cambiale, formation du bois et stress pédoclimatiques afin de améliorateurs est double : d'un côté, il est urgent de mieux comprendre les liens concevoir des génotypes mieux adaptés à leur environnement. […] Suite et fin du résumé dans la thèse. / Larix is a genus of high interest for plantation due to its fast growth rate and appreciated wood quality. Larch has a great potential to be introduced in middle and lowland afforestations as an alternative to Douglas-fir. One important question for breeders is adaptation to environment. It is particularly relevant in the context of climatic changes where a strong decrease of soil water availability during the growth period is predicted. Three larch species are used by foresters in France for plantation establishment, for which breeders must supply more adapted material: European and Japanese larch and their interspecific hybrid. The higher performance of hybrid larch in afforestations demonstrates its great potential. The tissue structure of annual rings reflects the response of cambium (cell division, elongation and thickening of cell walls) to environmental factors (climate, soil fertility, competition, etc), as well as physiological states and genetics. Then, wood density can be interpreted as the result of cambial activity and its variation as the response of the tree to environment. However, to gain a better understanding of this response and of the variation of the xylogenesis process itself, it is important to spot the timing of wood cell production. There are different approaches to study the timing of wood formation. We used a combination of wood microdensitometry and of pinning method as a new approach to study the dynamic of wood formation from a tree improvement point of view. This new approach allowed us to go beyond the static character of the microdensity profile (where density variation is related to distance) and to transform it into a dynamic profile (where density variation is related to time). Variation in phenology of cambial activity can be seen as a mechanism better adjusting trees to their environment. If this adjustment improves fitness, then the corresponding tree anatomical response to environment variation may be seen as an adaptive response. As breeders, our interest is twice: on one side, a better understanding of cambial phenology and of wood formation in relation to pedo-climatic factors and climatic stress is urgent for profiling genotypes better fitted to their environment. On the other side, exploitation design trees with better wood properties. In larch, phenotypic variation of variables through breeding of knowledge about phenology of wood formation will help to related to the dates of initiation and completion of the formation of different tissues and to the total duration of ring formation is low. Last and final summary in the thesis.

Vegetative propagation allows the amplification of selected genotypes for research, breeding, and commercial planting. However, efficient in vitro regeneration and genetic transformation remains a major obstacle to research and commercial application in many plant species. Our aims are to improve knowledge of gene regulatory circuits important to meristem organization, and to identify genes that might be useful for improving the efficiency of in vitro regeneration. In this thesis, we have approached these goals in two ways. First, we analyzed gene expression during poplar (Populus) regeneration using an AffymetrixGeneChip® array representing over 56,000 poplar transcripts. We have produced a catalog of regulated genes that can be used to inform studies of gene function and biotechnology. Second, we developed a GUS reporter system for monitoring meristem initiation using promoters of poplar homologs to the meristem-active regulatory genes WUSCHEL (WUS) and SHOOTMERISTEMLESS (STM). This provides plant materials whose developmental state can be assayed with improved speed and sensitivity. For the microarray study, we hybridized cDNAs derived from tissues of a female hybrid poplar clone (INRA 717-1 B4, Populus tremula x P. alba) at five sequential time points during organogenesis. Samples were taken from stems prior to callus induction, at 3 days and 5 days after callus induction, and at 3 and 8 days after the start of shoot induction. Approximately 15% of the monitored genes were significantly up-or down-regulated based on both Extraction and Analysis of Differentially Expressed Gene Expression (EDGE) and Linear Models for Microarray Data (LIMMA, FDR<0.01). Of these, over 3,000 genes had a 5-fold or greater change in expression. We found a very strong and rapid change in gene expression at the first time point after callus induction, prior to detectable morphological changes. Subsequent changes in gene expression at later regeneration stages were more than an order of magnitude smaller. A total of 588 transcription factors that were distributed in 45 gene families were differentially regulated. Genes that showed strong differential expression encoded proteins active in auxin and cytokinin signaling, cell division, and plastid development. When compared with data on in vitro callogenesis from root explants in Arabidopsis, 25% (1,260) of up-regulated and 22% (748) of down- regulated genes were in common with the genes that we found regulated in poplar during callus induction. When ~3kb of the 5' flanking regions of close homologs were used to drive expression of the GUSPlus gene, 50 to 60% of the transgenic events showed expression in apical and axillary meristems. However, expression was also common in other organs, including in leaf veins (40% and 46% of WUS and STM transgenic events, respectively) and hydathodes (56% of WUS transgenic events). Histochemical GUS staining of explants during callogenesis and shoot regeneration using in vitro stems as explants showed that expression was detectable prior to visible shoot development, starting 3 to 15 days after explants were placed onto callus inducing medium. Based on microarray gene expression data, a paralog of poplar WUS was detectably up-regulated during shoot initiation, but the other paralog was not. Surprisingly, both paralogs of poplar STM were down-regulated 3- to 6-fold during early callus initiation, a possible consequence of its stronger expression in the secondary meristem (cambium) than in shoot tissues. We identified 15 to 35 copies of cytokinin response regulator binding motifs (ARR1AT) and one copy of the auxin response element (AuxRE) in both promoters. Several of the WUS and STM transgenic events produced should be useful for monitoring the timing and location of meristem development during natural and in vitro shoot regeneration. / Graduation date: 2008