Abstract Ascorbate peroxidases (APXs) are heme peroxidases involved in the control of hydrogen peroxide levels and signal transduction pathways related to development and stress responses. Here, a total of 238 APX, 30 APX-related (APX-R), and 34 APX-like (APX-L) genes were identified from 24 species from the Poaceae family. Phylogenetic analysis of APX indicated five distinct clades, equivalent to cytosolic (cAPX), peroxisomal (pAPX), mitochondrial (mitAPX), stromal (sAPX), and thylakoidal (tAPX) isoforms. Duplication events contributed to the expansion of this family and the divergence times. Different from other APX isoforms, the emergence of Poaceae mitAPXs occurred independently after eudicot and monocot divergence. Our results showed that the constitutive silencing of mitAPX genes is not viable in rice plants, suggesting that these isoforms are essential for rice regeneration or development. We also obtained rice plants silenced individually to sAPX isoforms, demonstrating that, different to plants double silenced to both sAPX and tAPX or single silenced to tAPX previously obtained, these plants do not show changes in the total APX activity and hydrogen peroxide content in the shoot. Among rice plants silenced to different isoforms, plants silenced to cAPX showed a higher decrease in total APX activity and an increase in hydrogen peroxide levels. These results suggest that the cAPXs are the main isoforms responsible for regulating hydrogen peroxide levels in the cell, whereas in the chloroplast, this role is provided mainly by the tAPX isoform. In addition to broadening our understanding of the core components of the antioxidant defense in Poaceae species, the present study also provides a platform for their functional characterization.

O objetivo deste trabalho foi estabelecer sistemas de multiplicação de plantas de cana-de-açúcar in vitro e avaliar sua utilização, como material inicial, para a indução de regeneração a partir de ápices caulinares. Três métodos de cultivo foram avaliados: cultura em meio semi-sólido, cultura líquida estacionária e cultura líquida sob agitação. A taxa de multiplicação mais elevada foi alcançada por meio da cultura líquida sob agitação. Ápices caulinares, excisados dessas plantas, apresentaram taxas de regeneração in vitro compatíveis com sua utilização em protocolos de transformação. Calos resistentes a PPT e GUS-positivos foram obtidos de explantes da variedade Chunnee com inoculação de Agrobacterium tumefaciens C58C1 (pMP90) (pDUBarA9). O protocolo estabelecido a partir de cultivo in vitro pode ser utilizado para a produção de plantas transgênicas de cana-de-açúcar, visando à realização de estudos de regulação da expressão gênica, assim como à introdução de características de interesse agronômico.

The objective of this work was to establish in vitro systems for sugarcane plant multiplication and for regeneration from shoot apices excised from these plants. Three methods were analyzed: culture on semi-solid medium and liquid culture with or without agitation. The highest regeneration rates were obtained from cultures in liquid-medium with agitation. Shoot tips derived from these plants presented regeneration rates suitable for utilization in transformation protocols. PPT-resistant and GUS positive calluses were obtained from explants of in vitro plants of Chunnee variety inoculated with Agrobacterium tumefaciens C58C1 (pMP90) (pDUBarA9). The established in vitro culture system can be applied for transgenic sugarcane production, aiming at gene regulation studies and introduction of agronomical traits.

The occurrence of quasi-repetitive glycine-rich peptides has been reported in different organisms. Glycine-rich regions are proposed to be involved in protein-protein interactions in some mammalian protein families. In plants, a set of glycine-rich proteins (GRPs) was characterized several years ago, and since then a wealth of new GRPs have been identified. GRPs may have very diverse sub-cellular localization and functions. The only common feature among all different GRPs is the presence of glycine-rich repeat domains. The expression of genes encoding GRPs is developmentally regulated, and also induced, in several plant genera, by physical, chemical and biological factors. In addition to the highly modulated expression, several GRPs also show tissue-specific localization. GRPs specifically expressed in xylem, phloem, epidermis, anther tapetum and roots have been described. In this paper, the structural and functional features of these proteins in Eucalyptus are summarized. Since this is the first description of GRPs in this species, particular emphasis has been given to the expression pattern of these genes by analyzing their abundance and prevalence in the different cDNA-libraries of the Eucalyptus Genome Sequencing Project Consortium (ForEST). The comparison of GRPs from Eucalyptus and other species is also discussed.

A embriogênese somática em berinjela foi induzida a partir de explantes cotiledonares cultivados em meio MS suplementado com 54 µM de ANA. A análise anatômica do início de formação e desenvolvimento de embriões somáticos foi realizada durante as primeiras quatro semanas. A formação de pré-embriões foi observada após o segundo dia de cultura, diretamente a partir de células perivasculares ou a partir de massas pré-embriogênicas derivadas de massas meristemáticas indeterminadas (IMMs) formadas a partir do tecido vascular. As IMMs também deram origem a primórdios radiculares após 10 dias de cultura. A origem dos embriões é discutida, assim como as similaridades entre a formação de embriões somáticos e raízes adventícias.

Somatic embryogenesis was induced from cotyledon explants of eggplant cultured on MS medium supplemented with 54 µM NAA. Anatomical analysis of somatic embryo initiation and development was performed during the first four weeks. Proembryo formation was observed after the second day of culture, directly from perivascular cells or via pro-embryogenic masses derived from indeterminate meristematic masses (IMMs) originated in the vascular tissue. Those IMMs also gave rise to root primordia after 10 days of culture. The origin of embryos is discussed as well as the similarities between somatic embryogenesis and adventitious root formation.

The expression of AtchitIV gene was analysed in Arabidopsis plants submitted to abiotic stresses. Transcript accumulation was detected in leaves in response to UV light exposure, exogenous salicylic acid administration and wounding. Transgenic Arabidopsis plants carrying AtchitIV promoter::gus fusion also showed differential expression of the reporter gene in response to these treatments. The AtchitIV expression was also analysed during Arabidopsis embryo development. GUS assay demonstrated AtchitIV promoter activation in zygotic embryos from torpedo stage up to full maturation. Promoter deletion analysis indicated that all the 5' cis-acting elements responsible for the specific tissue expression are located in a region of 1083 bp, adjacent to the start of transcription. A negative regulatory region located between portions -1083 and -600 was also observed.

Ligninas são compostos fenólicos encontrados nas paredes secundárias do sistema vascular vegetal e desempenham importantes papéis biológicos, reduzindo a permeabilidade da parede celular em relação à água, estando também envolvidas em mecanismos de defesa contra patógenos. A via metabólica da lignina e as enzimas envolvidas na sua síntese vem sendo caracterizadas nos últimos anos. Uma série de genes que codificam diferentes enzimas envolvidas na biossíntese da lignina foram identificados em diferentes espécies de plantas. A biossíntese de lignina está acoplada ao metabolismo dos fenilpropanóides, apresentando enzimas compartilhadas com outros processos metabólicos tais como fenilalanina amônio liase (PAL), cinnamato 4-hidroxilase (C4H) and ácido caféico O-metiltransferase (COMT) assim como enzimas específicas como cinamoil-CoA redutase (CCR) e cinamil álcool dehidrogenase (CAD). Em certos tipos de mutantes de milho e sorgo, um aumento na digestibilidade foi associado a uma redução no teor de lignina. Uma redução na atividade de CAD e COMT, importantes enzimas envolvidas na biossíntese de lignina vem sendo demonstrada. Estas observações tem motivado diferentes grupos de pesquisa a alterarem o conteúdo ou a composição da lignina em plantas modelo, assim como culturas de interesse econômico, através de engenharia genética. O principal objetivo prático destes projetos é uma otimização da utilização destas plantas levando a uma maior produção de papel e digestibilidade da ração animal. No presente trabalho foi realizado um inventário das seqüências de ESTs, que codificam enzimas envolvidas no metabolismo de lignina, presentes no Projeto Genoma de Cana-de-açúcar (SUCEST). A análise foi realizada com as enzimas chaves ferulato-5-hidroxilase (F5H), ácido caféico O-metiltransferase (COMT), cafeoil CoA O-metiltransferase (CCoAOMT), hidroxicinamato CoA ligase (4CL), cinamoil-CoA redutase (CCR) and cinamil álcool dehidrogenase (CAD). A análise comparativa destes genes com os descritos em outras espécies poderá servir para a identificação de marcadores moleculares em programas de melhoramento genético, assim como em projetos que visem a manipulação do metabolismo de lignina em cana-de-açúcar.

Lignins are phenolic polymers found in the secondary wall of plant conductive systems where they play an important role by reducing the permeability of the cell wall to water. Lignins are also responsible for the rigidity of the cell wall and are involved in mechanisms of resistance to pathogens. The metabolic routes and enzymes involved in synthesis of lignins have been largely characterized and representative genes that encode enzymes involved in these processes have been cloned from several plant species. The synthesis of lignins is liked to the general metabolism of the phenylpropanoids in plants, having enzymes (e.g. phenylalanine ammonia-lyase (PAL), cinnamate 4-hydroxylase (C4H) and caffeic acid O-methyltransferase (COMT)) common to other processes as well as specific enzymes such as cinnamoyl-CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD). Some maize and sorghum mutants, shown to have defective in CAD and/or COMT activity, are easier to digest because they have a reduced lignin content, something which has motivated different research groups to alter the lignin content and composition of model plants by genetic engineering try to improve, for example, the efficiency of paper pulping and digestibility. In the work reported in this paper, we have made an inventory of the sugarcane expressed sequence tag (EST) coding for enzymes involved in lignin metabolism which are present in the sugarcane EST genome project (SUCEST) database. Our analysis focused on the key enzymes ferulate-5-hydroxylase (F5H), caffeic acid O-methyltransferase (COMT), caffeoyl CoA O-methyltransferase (CCoAOMT), hydroxycinnamate CoA ligase (4CL), cinnamoyl-CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD). The comparative analysis of these genes with those described in other species could be used as molecular markers for breeding as well as for the manipulation of lignin metabolism in sugarcane.

Desde o isolamento da primeira proteína rica em glicina (GRP) em plantas, um grande número de novas GRPs vem sendo identificado. Seu padrão de expressão altamente específico, embora variado, em conjunto com as diferentes localizações sub-celulares de alguns dos tipos de GRPs, claramente indica que estas proteínas encontram-se envolvidas em diversos processos fisiológicos independentes. Embora ainda sem uma clara definição do papel de GRPs na célula vegetal, estudos realizados com estas proteínas têm resultado em novos e interessantes esclarecimentos da biologia celular e molecular de plantas. Promotores com regulação complexa, assim como distintos mecanismos de regulação da expressão gênica tem sido demonstrados. Novas vias de endereçamento de proteínas tais como a exportação de GRPs para diferentes tipos celulares, tem sido observados. Estes dados mostram que as GRPs podem constituir marcadores e/ou modelos interessantes para a compreensão de distintos aspectos da biologia vegetal. Neste trabalho, características estruturais e funcionais deste tipo de proteínas em cana-de-açúcar (Saccharum officinarum L.) foram analisadas. Uma vez que esta é a primeira descrição deste tipo de proteínas em cana-de-açúcar, especial atenção foi dada para o padrão de expressão destes genes, analisando-se a abundância e prevalência de cada um dos genes nas diferentes bibliotecas de cDNA do projeto Sequenciamento de ESTs de cana-de-açúcar (SUCEST). A comparação das GRPs de cana-de-açúcar com as GRPs descritas em outras espécies também será discutida.

Since the isolation of the first glycine-rich proteins (GRPs) in plants a wealth of new GRPs have been identified. The highly specific but diverse expression pattern of grp genes, taken together with the distinct sub-cellular localization of some GRP groups, clearly indicate that these proteins are involved in several independent physiological processes. Notwithstanding the absence of a clear definition of the role of GRPs in plant cells, studies conducted with these proteins have provided new and interesting insights into the molecular biology and cell biology of plants. Complexly regulated promoters and distinct mechanisms for the regulation of gene expression have been demonstrated and new protein targeting pathways, as well as the exportation of GRPs from different cell types have been discovered. These data show that GRPs can be useful as markers and/or models to understand distinct aspects of plant biology. In this paper, the structural and functional features of these proteins in sugarcane (Saccharum officinarum L.) are summarized. Since this is the first description of GRPs in sugarcane, special emphasis has been given to the expression pattern of these GRP genes by studying their abundance and prevalence in the different cDNA-libraries of the Sugarcane Expressed Sequence Tag (SUCEST) project . The comparison of sugarcane GRPs with GRPs from other species is also discussed.