Proteomic Analysis of Chloroplast-to-Chromoplast Transition in Tomato Reveals Metabolic Shifts Coupled with Disrupted Thylakoid Biogenesis Machinery and Elevated Energy-Production Components

A comparative proteomic approach was performed to identify differentially expressed proteins in plastids at three stages of tomato (Solanum lycopersicum) fruit ripening (mature-green, breaker, red). Stringent curation and processing of the data from three independent replicates identified 1,932 proteins among which 1,529 were quantified by spectral counting. The quantification procedures have been subsequently validated by immunoblot analysis of six proteins representative of distinct metabolic or regulatory pathways. Among the main features of the chloroplast-to-chromoplast transition revealed by the study, chromoplastogenesis appears to be associated with major metabolic shifts: (1) strong decrease in abundance of proteins of light reactions (photosynthesis, Calvin cycle, photorespiration) and carbohydrate metabolism (starch synthesis/degradation), mostly between breaker and red stages and (2) increase in terpenoid biosynthesis (including carotenoids) and stress-response proteins (ascorbate-glutathione cycle, abiotic stress, redox, heat shock). These metabolic shifts are preceded by the accumulation of plastid-encoded acetyl Coenzyme A carboxylase D proteins accounting for the generation of a storage matrix that will accumulate carotenoids. Of particular note is the high abundance of proteins involved in providing energy and in metabolites import. Structural differentiation of the chromoplast is characterized by a sharp and continuous decrease of thylakoid proteins whereas envelope and stroma proteins remain remarkably stable. This is coincident with the disruption of the machinery for thylakoids and photosystem biogenesis (vesicular trafficking, provision of material for thylakoid biosynthesis, photosystems assembly) and the loss of the plastid division machinery. Altogether, the data provide new insights on the chromoplast differentiation process while enriching our knowledge of the plant plastid proteome.

Desempenho do cultivo da berinjela em plantio direto submetida a diferentes lâminas de irrigação

Este estudo foi conduzido no município de Seropédica, RJ, com o objetivo de se determinar, em cultivo orgânico e sistema de plantio direto, a produtividade da cultura da berinjela sob diferentes lâminas de irrigação e sistemas de cultivo (consorciada com leguminosa e solteira). O delineamento experimental adotado foi o de blocos ao acaso, no esquema de parcela subdividida com quatro repetições caracterizando, na parcela, os tratamentos equivalentes à lâmina de irrigação (40; 70; 100 e 120% ETc) e, na subparcela, os sistemas de cultivo consorciado com feijão caupi e solteiro. Ambos os sistemas de cultivo não influenciaram a produtividade final da berinjela; no entanto, considerando as diferentes lâminas, a maior produtividade comercial foi de 65,41 Mg ha-1, obtida para uma lâmina total de 690,04 mm (106,8% ETc). A menor lâmina aplicada resultou em qualidade inferior dos frutos em relação às maiores lâminas, sendo a taxa de descarte dos frutos de 3 e 14%, respectivamente, para a maior e a menor lâmina.

Composição, diversidade e distribuição geográfica de plantas vasculares de uma Floresta Ombrófila Densa Atlântica do Sudeste do Brasil

In the Montane and Submontane Rain Forest of the Carlos Botelho State Park - PECB (ca. 37,000 ha) the composition, richness and geographical distribution of native, vascular forest species was evaluated. The analysis of 1143 species of 140 families supported the pattern found for other forests of Eastern Brazil, showing high species richness of Myrtaceae (85 species), Orchidaceae (81), Fabaceae (57), Asteraceae, Melastomataceae (54), Lauraceae (53), Rubiaceae (51), Bromeliaceae (43), Piperaceae (30) and Solanaceae (25), besides ferns (123). The most species-rich genera were Eugenia (34), Ocotea (26), Leandra, Myrcia, Vriesea (18), Piper, Solanum (16), Miconia (14), Mollinedia (13), and Peperomia (12). The richness and composition varied greatly among life forms, as well as the number of families represented in each one of them (only Rubiaceae had species in all life forms, except parasites). Trees had the largest contribution of total richness (39.1%), a value that represented more than 20% of the species listed for the whole Atlantic Forest of Southeastern Brazil. Trees were followed by epiphytes (22.4%), herbs (18.4%), shrubs (10.1%), lianas (9.1%), and parasites (0.9%). The overall richness and composition of life forms was quite close to other neotropical forests (e.g. high contribution of ferns among epiphytes), although some life forms remain undersampled in the PECB (mainly herbs, lianas and epiphytes). The occurrence of species endemic to the Atlantic Forest was pronounced (65%), with a predominance of species restricted to the Southern Atlantic Forest (43%). Pantropical species were rare (2%), being more common among ferns. Myrtaceae and Melastomataceae were the families with greater number and proportion of endemic species.

Radiation of the Tnt1 retrotransposon superfamily in three Solanaceae genera

Abstract Background Tnt1 was the first active plant retrotransposon identified in tobacco after nitrate reductase gene disruption. The Tnt1 superfamily comprises elements from Nicotiana (Tnt1 and Tto1) and Lycopersicon (Retrolyc1 and Tlc1) species. The study presented here was conducted to characterise Tnt1-related sequences in 20 wild species of Solanum and five cultivars of Solanum tuberosum. Results Tnt1-related sequences were amplified from total genomic DNA using a PCR-based approach. Purified fragments were cloned and sequenced, and clustering analysis revealed three groups that differ in their U3 region. Using a network approach with a total of 453 non-redundant sequences isolated from Solanum (197), Nicotiana (140) and Lycopersicon (116) species, it is demonstrated that the Tnt1 superfamily can be treated as a population to resolve previous phylogenetic multifurcations. The resulting RNAseH network revealed that sequences group according to the Solanaceae genus, supporting a strong association with the host genome, whereas tracing the U3 region sequence association characterises the modular evolutionary pattern within the Tnt1 superfamily. Within each genus, and irrespective of species, nearly 20% of Tnt1 sequences analysed are identical, indicative of being part of an active copy. The network approach enabled the identification of putative "master" sequences and provided evidence that within a genus these master sequences are associated with distinct U3 regions. Conclusion The results presented here support the hypothesis that the Tnt1 superfamily was present early in the evolution of Solanaceae. The evidence also suggests that the RNAseH region of Tnt1 became fixed at the host genus level whereas, within each genus, propagation was ensured by the diversification of the U3 region. Different selection pressures seemed to have acted on the U3 and RNAseH modules of ancestral Tnt1 elements, probably due to the distinct functions of these regions in the retrotransposon life cycle, resulting in both co evolution and adaptation of the element population with its host.

Biochemical and histological characterization of tomato mutants

Biochemical responses inherent to antioxidant systems as well morphological and anatomical properties of photomorphogenic, hormonal and developmental tomato mutants were investigated. Compared to the non-mutant Micro-Tom (MT), we observed that the malondialdehyde (MDA) content was enhanced in the diageotropica (dgt) and lutescent (l) mutants, whilst the highest levels of hydrogen peroxide (H2O2) were observed in high pigment 1 (hp1) and aurea (au) mutants. The analyses of antioxidant enzymes revealed that all mutants exhibited reduced catalase (CAT) activity when compared to MT. Guaiacol peroxidase (GPOX) was enhanced in both sitiens (sit) and notabilis (not) mutants, whereas in not mutant there was an increase in ascorbate peroxidase (APX). Based on PAGE analysis, the activities of glutathione reductase (GR) isoforms III, IV, V and VI were increased in l leaves, while the activity of superoxide dismutase (SOD) isoform III was reduced in leaves of sit, epi, Never ripe (Nr) and green flesh (gf) mutants. Microscopic analyses revealed that hp1 and au showed an increase in leaf intercellular spaces, whereas sit exhibited a decrease. The au and hp1 mutants also exhibited a decreased in the number of leaf trichomes. The characterization of these mutants is essential for their future use in plant development and ecophysiology studies, such as abiotic and biotic stresses on the oxidative metabolism.

Relationship between eggplant seed morphology and germination

Structural differences such as abnormalities, damage and free spaces in seeds may affect germination. The aim of this study was to study the relationship between eggplant seed morphology and seed germination. Ten seed lots of the eggplant cultivar Embu were evaluated by X-ray image analysis and the germination test. Seed image analysis was performed by Image Pro Plus® software and the whole seed area and free space between the embryo and endosperm were measured. The internal seed area filled by the embryo and endosperm was calculated from the difference between the whole seed and free space areas. Based on these results and visual seed analysis, seeds were classified into three categories and information on germination was obtained for each one. X-ray image analysis provides a perfect view of the internal seed parts and for seed morphology studies. An increase in seed area filled by the endosperm and embryo does not improve seed germination. Mechanical seed damage and deteriorated tissues can adversely affect seed germination.

Desempenho de híbridos de tomate de crescimento indeterminado em função de sistemas de condução de plantas

O presente trabalho teve como objetivo avaliar dois sistemas de tutoramento e condução de plantas na produtividade e seus componentes em híbridos de tomateiro dos segmentos Salada Longa Vida e Santa Cruz de crescimento indeterminado, visando ao consumo in natura. Adotou-se o delineamento em blocos ao acaso com os tratamentos em esquema fatorial 5x2, quatro repetições, sendo avaliados cinco híbridos comerciais, três pertencentes ao grupo Salada Longa Vida ('Alambra', 'Paron' e 'Forty') e dois do grupo Santa Cruz ('Débora Pto' e 'Ellus'), em dois sistemas de condução de plantas (bambu e fitilho). O experimento foi conduzido em condições de campo, em Itatiba (SP), de julho de 2007 a janeiro de 2008. Foram consideradas todas as plantas de cada parcela e avaliaram-se a produtividade total, produtividade comercial, produção por planta, massa média de frutos, número total de frutos, número de frutos comercializáveis, número de frutos das classes '3A', '2A' e '1A', número de frutos manchado e com lóculo aberto. Não houve interação entre cultivar e sistema de condução de planta para maioria das características avaliadas, exceto número total de frutos, número de frutos comercializáveis e número de frutos da classe '2A'. O sistema de condução de plantas com uma haste tutorada com fitilho proporcionou incremento na massa média de frutos e número de frutos da classe '3A', e decréscimo do número de frutos da classe '1A', sem reduzir a produtividade total e comercial quando comparado ao método de condução tradicional no bambu com duas hastes por planta.

Diallel cross among fresh market tomato inbreeding lines

The general combining ability (GCA), specific combining ability (SCA), and heterosis were studied in a complete diallel cross among fresh market tomato breeding lines with reciprocal excluded. Fifteen genotypes (five parents and ten hybrids) were tested using a randomized complete block design, with three replications, and the experiments were conducted in Itatiba, São Paulo state, Brazil, in 2005/06. The yield components evaluated were fruit yield per plant (FP), fruit number per plant (FN), average fruit weight (FW); cluster number per plant (CN); fruit number per cluster (FC), fruit wall thickness (FT) and number of locules per fruit (NL). Fruit quality components evaluated were total soluble solids (SS); total titratable acidity (TA); SS/TA ratio, fruit length (FL); fruit width (WI); length to width ratio (FL/WI). The data for each trait was first subjected to analysis of variance. Griffing's method 2, model 1 was employed to estimate the general (GCA) and specific (SCA) combining abilities. Parental and hybrid data for each trait were used to estimate of mid-parent heterosis. For plant fruit yield, IAC-2 was the best parental line with the highest GCA followed by IAC-4 and IAC-1 lines. The hybrids IAC-1 x IAC-2, IAC-1 x IAC-4 and IAC-2 x IAC-4 showed the highest effects of SCA. High heterotic responses were found for fruit yield and plant fruit number with values up to 49.72% and 47.19%, respectively. The best hybrids for fruit yield and plant fruit number were IAC-1 x IAC-2, IAC-1 x IAC-4 and IAC-2 x IAC-5, for fruit yield and plant fruit number, the main yield components.

Correlations between yield and fruit quality characteristics of fresh market tomatoes

Genotypic, phenotypic and environmental correlations were estimated for all possible pairs among eleven characters of tomatoes. Fifteen treatments including five parents and ten hybrids of Instituto Agronômico (IAC) tomato breeding program were evaluated using a randomized complete block experimental design, with tree replications in Itatiba, São Paulo state, Brazil, during 2005/2006. The following traits were evaluated: fruit yield per plant (FP), fruit number per plant (FN), average fruit weight (FW), cluster number per plant (CN), fruit number per cluster (FC), number of locules per fruit (NL), fruit length (FL), fruit width (WI), fruit wall thickness (FT), total soluble solids (SS), and total titratable acidity (TA). The genotypic (rG), phenotypic (rF) and environmental correlations (rA) for two pairs of plant traits were estimated using the Genes© program. High similarity was found among the estimates of genotypic and phenotypic correlations. Positive and high phenotypic and genotypic correlations were observed between FP and the traits FN, FW and FT, and these associations contributed for yield increasing. FW and FT contributed to yield increase and should be considered together as primary yield components in tomato. Positive values of the genotypic and phenotypic correlations revealed that FP influenced FN with high direct effect and significant positive correlation. These traits may be included as the main selection criteria for tomato yield improvement.

Base genética do hábito de crescimento e florescimento em tomateiro e sua importância na agricultura

Variedades ou híbridos de tomateiro utilizados para produção de molhos e ketchups costumam ter hábito determinado, enquanto a maioria dos genótipos utilizados na produção para mesa (consumo in natura) possuem hábito indeterminado. Além de influenciar no manejo da cultura, o hábito de crescimento pode alterar parâmetros de produtividade, como o teor de sólidos solúveis totais (SST) nos frutos. O hábito de crescimento é controlado principalmente pelo gene SELF-PRUNING (SP), que é um dos componentes de uma pequena família gênica da qual faz parte também o gene SINGLE FLOWER TRUSS (SFT), atualmente considerado um dos componentes do tão buscado "florígeno". O entendimento da função bioquímica e o efeito fisiológico de tais genes em interação com o ambiente e outros genes (epistasia) possibilita a manipulação de parâmetros como precocidade e SST. Além disso, fornece subsídios para compreender a base genética do crescimento semideterminado, que combina vantagens do hábito determinado e indeterminado, podendo ser usado pelos melhoristas de plantas para o desenvolvimento de novas cultivares.

Physiological quality of osmoprimed eggplant seeds

Eggplant seeds germination can be slow and uneven, justifying the use of pre-germinative treatments to improve the performance of seed lots. One option of treatment is the controlled hydration of seeds by priming. In this way, this study aimed to evaluate the performance of eggplant seeds cv. Embu submitted to different methodologies of priming. The seeds used in the experiment were stored in cold chamber (15º C and 55% RH) in paper bags. The research was carried out at Central Laboratory of Seeds/UFLA. The seeds were submitted to the priming in aerated solutions varying the following factors: temperature (15º C and 25º C), time (24, 48 and 72 hours) and solution (water, PEG, KNO3 and PEG+KNO3). Seeds were washed in running water and dried at 30º C, until the return to the initial moisture content, around 10%. The variables analyzed were percentage of germination, percentage of emergence, speed index of emergence and electrical conductivity. The treatments were arranged in a completely randomized design, according to a factorial arrangement 2x3x4+1 (control - seeds without priming). The results showed that priming improves the vigour of eggplant seeds with no effect on viability; the priming in water or KNO3 is efficient to improve the seed vigour and priming in water or KNO3 may use temperature of 15º C or 25º C for 24, 48 or 72 hours.

Microalgae bio-stimulating effect on traditional tomato agriculture

Máster en Oceanografía

Studies on biotic and abiotic elicitors inducing defense responses in tomato

Tomato (Lycopersicon esculentum Mill., Solanum lycopersicon L.) is one of the most popular vegetable throughout the world, and the importance of its cultivation is threatened by a wide array of pathogens. In the last twenty years this plant has been successfully used as a model plant to investigate the induction of defense pathways after exposure to fungal, bacterial and abiotic molecules, showing triggering of different mechanisms of resistance. Understanding these mechanisms in order to improve crop protection is a main goal for Plant Pathology. The aim of this study was to search for general or race-specific molecules able to determine in Solanum lycopersicon immune responses attributable to the main systems of plant defense: non-host, host-specific and induced resistance. Exopolysaccharides extracted by three fungal species (Aureobasidium pullulans, Cryphonectria parasitica and Epicoccum purpurascens), were able to induce transcription of pathogenesis-related (PR) proteins and accumulation of enzymes related to defense in tomato plants cv Money Maker,using the chemical inducer Bion® as a positive control. During the thesis, several Pseudomonas spp. strains were also isolated and tested for their antimicrobial activity and ability to produce antibiotics. Using as a positive control jasmonic acid, one of the selected strain was shown to induce a form of systemic resistance in tomato. Transcription of PRs and reduction of disease severity against the leaf pathogen Pseduomonas syringae pv. tomato was determined in tomato plants cv Money Maker and cv Perfect Peel, ensuring no direct contact between the selected rhizobacteria and the aerial part of the plant. To conclude this work, race-specific resistance of tomato against the leaf mold Cladosporium fulvum is also deepened, describing the project followed at the Phytopathology Laboratory of Wageningen (NL) in 2007, dealing with localization of a specific R-Avr interaction in transfected tomato protoplast cultures through fluorescence microscopy.

Leaf Asymmetry as a Developmental Constraint Imposed by Auxin-Dependent Phyllotactic Patterning

In a majority of species, leaf development is thought to proceed in a bilaterally symmetric fashion without systematic asymmetries. This is despite the left and right sides of an initiating primordium occupying niches that differ in their distance from sinks and sources of auxin. Here, we revisit an existing model of auxin transport sufficient to recreate spiral phyllotactic patterns and find previously overlooked asymmetries between auxin distribution and the centers of leaf primordia. We show that it is the direction of the phyllotactic spiral that determines the side of the leaf these asymmetries fall on. We empirically confirm the presence of an asymmetric auxin response using a DR5 reporter and observe morphological asymmetries in young leaf primordia. Notably, these morphological asymmetries persist in mature leaves, and we observe left-right asymmetries in the superficially bilaterally symmetric leaves of tomato (Solanum lycopersicum) and Arabidopsis thaliana that are consistent with modeled predictions. We further demonstrate that auxin application to a single side of a leaf primordium is sufficient to recapitulate the asymmetries we observe. Our results provide a framework to study a previously overlooked developmental axis and provide insights into the developmental constraints imposed upon leaf morphology by auxin-dependent phyllotactic patterning.

Regulation of woody plant development by gibberellin catabolic and signaling genes

Gibberellin (GA) is a growth promoting hormone implicated in regulating a diversity of plant processes. This dissertation examines the role of GA metabolic and signaling genes in woody plant growth and development. Transgenic modifications, expression analysis, physiological/biochemical assays, biometric measurements and histological analysis were used to understand the regulatory roles these genes play in the model woody plant, Populus. Our results highlight the importance of GA regulatory genes in woody perennial growth, including: phenology, wood formation, phenotypic plasticity, and growth/survival under field conditions. We characterize two putative Populus orthologs of the SHORT INTERNODES (SHI) gene from Arabidopsis, a negative regulator of GA signaling. RNAi-mediated suppression of Populus SHI-like genes increased several growth-related traits, including extent of xylem proliferation, in a dose-dependent manner. Three Populus genes, sharing sequence homology to the positive regulator of GA signaling gene PHOTOPERIOD-RESPONSIVE 1 (PHOR1) from Solanum, are up-regulated in GA-deficient and insensitive plants suggesting a conserved role in GA signaling. We demonstrate that Populus PHOR1-like genes have overlapping and divergent function(s). Two PHOR1-like genes are highly expressed in roots, predominantly affect root growth (e.g., morphology, starch quantity and gravitropism), and induced by short-days (SD). The other PHOR1-like gene is ubiquitously expressed with a generalized function in root and shoot development. The effects of GA catabolic and signaling genes on important traits (e.g., adaptive and productivity traits) were studied in a multi-year field trial. Transgenics overexpressing GA 2-oxidase (GA2ox) and DELLA genes showed tremendous variation in growth, form, foliage, and phenology (i.e., vegetative and reproductive). Observed gradients in trait modifications were correlated to transgene expression levels, in a manner suggesting a dose-dependent relationship. We explore GA2ox and DELLA genes involvement in mediating growth responses to immediate short-term drought stress, and SD photoperiods, signaling prolonged periods of stress (e.g., winter bud dormancy). GA2ox and DELLA genes show substantial up-regulation in response to drought and SDs. Transgenics overexpressing homologs of these genes subjected to drought and SD photoperiods show hypersensitive growth restraint and increased stress resistances. These results suggest growth cessation (i.e., dormancy) in response to adverse conditions is mediated by GA regulatory genes.