Publications by authors named "Wagner C Otoni"

19 Publications

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Deleterious effects of Pfaffia glomerata (Spreng.) Pedersen hydroalcoholic extract on the seminiferous epithelium of adult Balb/c mice.

Int J Exp Pathol 2020 10 1;101(5):183-191. Epub 2020 Sep 1.

Departmento de Biologia Geral, Universidade Federal de Viçosa, Viçosa, Brasil.

Several plant species such as Pfaffia glomerata are widely used in traditional Brazilian medicine as stimulants and aphrodisiacs. In this regard, the aim of our study was to explore the effects of the long-term intake of the hydro-alcoholic root extract of P glomerata on the germ and somatic cells within the seminiferous tubules in adult Balb/c mice. The experimental groups were placed as: controls (water and DMSO), and treated with 300 and 400 mg/kg of the root extract. The number of germ and somatic cells, the proportion of pathological seminiferous tubules, and the germ cell apoptotic levels were evaluated. The volume and proportion of the seminiferous epithelium was decreased after the extract intake due to the increased germ cell apoptotic levels. Vacuolization of Sertoli cell cytoplasm was observed widely in pathological tubules, along with fully disorganized epithelia, showing multinucleated cells, which lead to decreased daily sperm production. Taken together, our results indicate that long-term intake of the P glomerata caused deleterious effects on spermatogenesis by inducing apoptosis and altering the seminiferous tubule's epithelial dynamics.
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http://dx.doi.org/10.1111/iep.12363DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7495768PMC
October 2020

Compatibility in pollen-pistil interaction of interspecific crossings with Passiflora spp.

An Acad Bras Cienc 2020 27;92 Suppl 1:e20180062. Epub 2020 Jul 27.

Departamento de Biologia Vegetal, Universidade Federal de Viçosa, Viçosa, MG, Brazil.

Intraspecies or interspecies crossings transfer relevant alleles between plants. However, some interspecies crossings involving Passiflora species impede ovule fertilization and the viable development of seeds. Thus, the purpose of this study was to verify the viability of interspecific crossings and monitor pollen tube development. The experiment had six species of Passiflora in the reciprocal crossings. Histochemical tests aimed to evaluate the percentage of intraspecies or interspecies crossings that resulted in fruit development and pollen tube development. Ovule fertilization and fruit development occurred in determined directions of crossings when controlling the female or male genitor, but only one case of reciprocal crossing had success. In crossings with no fruit development, histological analysis showed that some callus developed in the stigma and style, confirming unilateral and interspecies incompatibility in the genus Passiflora to some species and some directions of crossings.
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http://dx.doi.org/10.1590/0001-3765202020180062DOI Listing
September 2020

Somatic embryogenesis in the commercial papaya hybrid UENF/Caliman 01 relying on plantlet production from sexed adult hermaphrodite donor plants.

An Acad Bras Cienc 2019 Aug 19;91(3):e20180504. Epub 2019 Aug 19.

Departamento de Ciências Florestais e da Madeira, CCAE/UFES. Avenida Governador Lindemberg, 316, Centro, 29550-000 Jerônimo Monteiro, ES, Brazil.

Somatic embryogenesis from explants from hermaphrodite papaya mother plants is an alternative for the production of true-to-type plants without the need for sexing. This study aimed to analyze hormonal and osmotic inducers in different somatic embryogenesis stages in the commercial hermaphrodite hybrid papaya UENF/Caliman 01. Leaf disks from in vitro shoots originated from ex vitro hermaphrodite plants were cultured in induction medium supplemented with different concentrations of 2,4-D (6, 9, 12, 15, and 18 μM) and 4-CPA (19, 22, 25, 28, and 31 μM). After 90 days, the formation of somatic embryos was verified. The 2,4-D induced the formation of light brown calli with low frequency (20%) of somatic embryogenesis. However, 4-CPA (25 μM) induced 96% of embryogenic calli, which were transferred to maturation medium (MM) and cultured for 30 days. The MM contained ABA (0.5 μM) and AC (15 g L-1) and produced 36.6 somatic embryos callus-1, mainly on cotyledonary stage. Cotyledonary embryos were transferred to germination medium supplemented with gibberellic acid (GA3) (0.0, 1.44, 2.88, and 4.32 µM), and the conversion into plantlets was enhanced with GA3 at 2.88 µM.
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http://dx.doi.org/10.1590/0001-3765201920180504DOI Listing
August 2019

Leaf heteroblasty in Passiflora edulis as revealed by metabolic profiling and expression analyses of the microRNAs miR156 and miR172.

Ann Bot 2019 07;123(7):1191-1203

Departamento de Biologia Vegetal/Instituto de Biotecnologia Aplicada a Agropecuária (BIOAGRO), Universidade Federal de Viçosa, Viçosa, Minas Gerais, Brazil.

Background And Aims: Juvenile-to-adult phase transition is marked by changes in leaf morphology, mostly due to the temporal development of the shoot apical meristem, a phenomenon known as heteroblasty. Sugars and microRNA-controlled modules are components of the heteroblastic process in Arabidopsis thaliana leaves. However, our understanding about their roles during phase-changing in other species, such as Passiflora edulis, remains limited. Unlike Arabidopsis, P. edulis (a semi-woody perennial climbing vine) undergoes remarkable changes in leaf morphology throughout juvenile-to-adult transition. Nonetheless, the underlying molecular mechanisms are unknown.

Methods: Here we evaluated the molecular mechanisms underlying the heteroblastic process by analysing the temporal expression of microRNAs and targets in leaves as well as the leaf metabolome during P. edulis development.

Key Results: Metabolic profiling revealed a unique composition of metabolites associated with leaf heteroblasty. Increasing levels of glucose and α-trehalose were observed during juvenile-to-adult phase transition. Accumulation of microRNA156 (miR156) correlated with juvenile leaf traits, whilst miR172 transcript accumulation was associated with leaf adult traits. Importantly, glucose may mediate adult leaf characteristics during de novo shoot organogenesis by modulating miR156-targeted PeSPL9 expression levels at early stages of shoot development.

Conclusions: Altogether, our results suggest that specific sugars may act as co-regulators, along with two microRNAs, leading to leaf morphological modifications throughout juvenile-to-adult phase transition in P. edulis.
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http://dx.doi.org/10.1093/aob/mcz025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6612941PMC
July 2019

The effects of an osmoregulator, carbohydrates and polyol on maturation and germination of 'Golden THB' papaya somatic embryos.

An Acad Bras Cienc 2018 Oct-Dec;90(4):3433-3447. Epub 2018 Oct 25.

Departamento de Ciências Florestais e da Madeira, Centro de Ciências Agrárias e Engenharias, Universidade Federal do Espírito Santo, Avenida Governador Lindemberg, 316, Centro, 29550-000 Jerônimo Monteiro, ES, Brazil.

This study evaluated the effect of osmoregulators and carbohydrates on the maturation and germination of somatic embryos of papaya 'Golden THB'. Cotyledon explants from papaya seedlings germinated in vitro on basal MS medium were cultured on somatic embryogenesis induction medium (IM) containing MS salts, myo-inositol, sucrose, agar and p-chlorophenoxyacetic acid. After 50 days, embryogenic calli were transferred onto maturation media (MM) for 45 additional days. For experiment 1, a MS-based medium supplemented with abscisic acid, activated charcoal and concentrations of PEG 6000 (0; 40; 50; 60 and 70 g L-1) was used, whereas for experiment 2 malt extract concentrations (0; 0.1; 0.2; 0.3 and 0.4 g L-1) were assessed. The normal cotyledonary somatic embryos produced in experiment 2 were transferred to the germination medium (GM). The GM consisted of full-strength MS medium, sucrose, agar and was supplemented with myo-inositol at varying concentrations (0; 0.275; 0.55 and 0.825 mM). The PEG concentrations tested impaired the maturation of 'Golden THB' papaya somatic embryos. The MM, supplemented with malt extract at 0.153 g L-1, promoted the greatest development of normal somatic embryos (18.28 SE calli-1), that is, two cotyledonary leaves produced 36.56 SE calli-1. The supplementation with 0.45 mM myo-inositol provided the highest germination percentage (47.42%) and conversion to emblings.
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http://dx.doi.org/10.1590/0001-3765201820171035DOI Listing
February 2019

Topological Data Analysis as a Morphometric Method: Using Persistent Homology to Demarcate a Leaf Morphospace.

Front Plant Sci 2018 25;9:553. Epub 2018 Apr 25.

Department of Ecology and Evolutionary Biology, Yale University, New Haven, CT, United States.

Current morphometric methods that comprehensively measure shape cannot compare the disparate leaf shapes found in seed plants and are sensitive to processing artifacts. We explore the use of persistent homology, a topological method applied as a filtration across simplicial complexes (or more simply, a method to measure topological features of spaces across different spatial resolutions), to overcome these limitations. The described method isolates subsets of shape features and measures the spatial relationship of neighboring pixel densities in a shape. We apply the method to the analysis of 182,707 leaves, both published and unpublished, representing 141 plant families collected from 75 sites throughout the world. By measuring leaves from throughout the seed plants using persistent homology, a defined morphospace comparing all leaves is demarcated. Clear differences in shape between major phylogenetic groups are detected and estimates of leaf shape diversity within plant families are made. The approach predicts plant family above chance. The application of a persistent homology method, using topological features, to measure leaf shape allows for a unified morphometric framework to measure plant form, including shapes, textures, patterns, and branching architectures.
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http://dx.doi.org/10.3389/fpls.2018.00553DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5996898PMC
April 2018

Selection and validation of reference genes for quantitative gene expression analyses in various tissues and seeds at different developmental stages in L.

Physiol Mol Biol Plants 2018 May 3;24(3):369-378. Epub 2018 Apr 3.

1Departamento de Ciências Biológicas, Universidade Estadual de Santa Cruz, Ilhéus, Bahia 45662-900 Brazil.

L., popularly known as annatto, produces several secondary metabolites of pharmaceutical and industrial interest, including bixin, whose molecular basis of biosynthesis remain to be determined. Gene expression analysis by quantitative real-time PCR (qPCR) is an important tool to advance such knowledge. However, correct interpretation of qPCR data requires the use of suitable reference genes in order to reduce experimental variations. In the present study, we have selected four different candidates for reference genes in , coding for 40S ribosomal protein S9 (RPS9), histone H4 (H4), 60S ribosomal protein L38 (RPL38) and 18S ribosomal RNA (18SrRNA). Their expression stabilities in different tissues (e.g. flower buds, flowers, leaves and seeds at different developmental stages) were analyzed using five statistical tools (NormFinder, geNorm, BestKeeper, ΔCt method and RefFinder). The results indicated that is the most stable gene in different tissues and stages of seed development and is the most unstable among the analyzed genes. In order to validate the candidate reference genes, we have analyzed the relative expression of a target gene coding for carotenoid cleavage dioxygenase 1 (CCD1) using the stable and the least stable gene, , for normalization of the qPCR data. The results demonstrated significant differences in the interpretation of the gene expression data, depending on the reference gene used, reinforcing the importance of the correct selection of reference genes for normalization.
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http://dx.doi.org/10.1007/s12298-018-0528-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5911269PMC
May 2018

Somatic embryogenesis in Carica papaya as affected by auxins and explants, and morphoanatomical-related aspects.

An Acad Bras Cienc 2018 Jan-Mar;90(1):385-400. Epub 2018 Feb 1.

Universidade Federal do Espírito Santo, Departamento de Ciências Florestais e da Madeira, Avenida Governador Lindemberg, 360, Centro, 29550-000 Jerônimo Monteiro, ES, Brazil.

The aim of this study was to evaluate somatic embryogenesis in juvenile explants of the THB papaya cultivar. Apical shoots and cotyledonary leaves were inoculated in an induction medium composed of different concentrations of 2,4-D (6, 9, 12, 15 and 18 µM) or 4-CPA (19, 22, 25, 28 and 31 µM). The embryogenic calluses were transferred to a maturation medium for 30 days. Histological analysis were done during the induction and scanning electron microscopy after maturing. For both types of auxin, embryogenesis was achieved at higher frequencies with cotyledonary leaves incubated in induction medium than with apical shoots; except for callogenesis. The early-stage embryos (e.g., globular or heart-shape) predominated. Among the auxins, best results were observed in cotyledonary leaves induced with 4-CPA (25 µM). Histological analyses of the cotyledonary leaf-derived calluses confirmed that the somatic embryos (SEs) formed from parenchyma cells, predominantly differentiated via indirect and multicellular origin and infrequently via synchronized embryogenesis. The secondary embryogenesis was observed during induction and maturation phases in papaya THB cultivar. The combination of ABA (0.5 µM) and AC (15 g L-1) in maturation medium resulted in the highest somatic embryogenesis induction frequency (70 SEs callus-1) and the lowest percentage of early germination (4%).
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http://dx.doi.org/10.1590/0001-3765201820160252DOI Listing
May 2018

Divergent leaf shapes among species arise from a shared juvenile morphology.

Plant Direct 2017 Nov 6;1(5):e00028. Epub 2017 Nov 6.

Departamento de Biologia Vegetal/BIOAGRO Universidade Federal de Viçosa Viçosa MG Brasil.

Not only does leaf shape vary between species, but between sequential nodes of the vine. The profound changes in leaf shape within vines reflect the temporal development of the shoot apical meristem from which leaves are derived and patterned, a phenomenon known as heteroblasty. We perform a morphometric analysis of more than 3,300 leaves from 40 different species using two different methods: homologous landmarks and Elliptical Fourier Descriptors (EFDs). Changes in leaf shape across the vine are first quantified in allometric terms; that is, changes in the relative area of leaf subregions expressed in terms of overall leaf area. Shape is constrained to strict linear relationships as a function of size that vary between species. Statistical analysis of leaf shape, using landmarks and EFDs, reveals that species effects (regardless of node) are the strongest, followed by interaction effects between species and heteroblasty (i.e., species-specific patterns in leaf shape across nodes) and that heteroblasty effects across nodes (regardless of species) are negligible. The ability of different nodes to predictively discriminate species and the variability of landmark and EFD traits at each node is then analyzed. Heteroblastic trajectories, the changes in leaf shape between the first and last measured leaves in a vine, are then compared between species in a multivariate space. Leaf shape diversity among species is expressed in a heteroblastic-dependent manner, unique to each species. Leaf shape is constrained by linear, allometric relationships related to leaf size that vary between species. There is a strong species × heteroblasty interaction effect for leaf shape, suggesting that different leaf shapes between species arise through changes in shape across nodes specific to each species. The first leaves in the series are not only more like each other, but are also less variable across species. From this similar, shared leaf shape, subsequent leaves in the heteroblastic series follow divergent morphological trajectories. The disparate leaf shapes characteristic of species arise from a shared, juvenile morphology.
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http://dx.doi.org/10.1002/pld3.28DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6508542PMC
November 2017

Erratum to: Morphometric analysis of Passiflora leaves: the relationship between landmarks of the vasculature and elliptical Fourier descriptors of the blade.

Gigascience 2017 10;6(10)

Background: Leaf shape among Passiflora species is spectacularly diverse. Underlying this diversity in leaf shape are profound changes in the patterning of the primary vasculature and laminar outgrowth. Each of these aspects of leaf morphology-vasculature and blade-provides different insights into leaf patterning.

Results: Here, we morphometrically analyze >3300 leaves from 40 different Passiflora species collected sequentially across the vine. Each leaf is measured in two different ways: using 1) 15 homologous Procrustes-adjusted landmarks of the vasculature, sinuses, and lobes; and 2) Elliptical Fourier Descriptors (EFDs), which quantify the outline of the leaf. The ability of landmarks, EFDs, and both datasets together are compared to determine their relative ability to predict species and node position within the vine. Pairwise correlation of x and y landmark coordinates and EFD harmonic coefficients reveals close associations between traits and insights into the relationship between vasculature and blade patterning.

Conclusions: Landmarks, more reflective of the vasculature, and EFDs, more reflective of the blade contour, describe both similar and distinct features of leaf morphology. Landmarks and EFDs vary in ability to predict species identity and node position in the vine and exhibit a correlational structure (both within landmark or EFD traits and between the two data types) revealing constraints between vascular and blade patterning underlying natural variation in leaf morphology among Passiflora species.
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http://dx.doi.org/10.1093/gigascience/gix070DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5632300PMC
October 2017

Morphometric analysis of Passiflora leaves: the relationship between landmarks of the vasculature and elliptical Fourier descriptors of the blade.

Gigascience 2017 01;6(1):1-13

Departamento de Biologia Vegetal/BIOAGRO, Universidade Federal de Viçosa, Viçosa, MG, Brasil.

Background: Leaf shape among Passiflora species is spectacularly diverse. Underlying this diversity in leaf shape are profound changes in the patterning of the primary vasculature and laminar outgrowth. Each of these aspects of leaf morphology-vasculature and blade-provides different insights into leaf patterning.

Results: Here, we morphometrically analyze >3300 leaves from 40 different Passiflora species collected sequentially across the vine. Each leaf is measured in two different ways: using 1) 15 homologous Procrustes-adjusted landmarks of the vasculature, sinuses, and lobes; and 2) Elliptical Fourier Descriptors (EFDs), which quantify the outline of the leaf. The ability of landmarks, EFDs, and both datasets together are compared to determine their relative ability to predict species and node position within the vine. Pairwise correlation of x and y landmark coordinates and EFD harmonic coefficients reveals close associations between traits and insights into the relationship between vasculature and blade patterning.

Conclusions: Landmarks, more reflective of the vasculature, and EFDs, more reflective of the blade contour, describe both similar and distinct features of leaf morphology. Landmarks and EFDs vary in ability to predict species identity and node position in the vine and exhibit a correlational structure (both within landmark or EFD traits and between the two data types) revealing constraints between vascular and blade patterning underlying natural variation in leaf morphology among Passiflora species.
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http://dx.doi.org/10.1093/gigascience/giw008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5437945PMC
January 2017

Erratum to: Antigen production using heterologous expression of dengue virus-2 non structural protein 1 (NS1) in Nicotiana tabacum (Havana) for immunodiagnostic purposes.

Plant Cell Rep 2016 Oct;35(10):2205

Laboratory of Molecular Immunovirology, Department of General Biology, Federal University of Viçosa, Av. PH Rolfs, s/n Campus Universitário, Viçosa, MG, CEP 36570-000, Brazil.

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http://dx.doi.org/10.1007/s00299-016-2043-6DOI Listing
October 2016

Positive selection on the K domain of the AGAMOUS protein in the Zingiberales suggests a mechanism for the evolution of androecial morphology.

Evodevo 2015 8;6. Epub 2015 Apr 8.

Department of Plant and Microbial Biology, University of California, Berkeley, 111 Koshland Hall, Berkeley, CA 94720 USA ; Department of Integrative Biology and the University and Jepson Herbaria, University of California, Berkeley, Berkeley, CA 94720 USA.

Background: The ABC model of flower development describes the molecular basis for specification of floral organ identity in model eudicots such as Arabidopsis and Antirrhinum. According to this model, expression of C-class genes is linked to stamen and gynoecium organ identity. The Zingiberales is an order of tropical monocots in which the evolution of floral morphology is characterized by a marked increase in petaloidy in the androecium. Petaloidy is a derived characteristic of the ginger families and seems to have arisen in the common ancestor of the ginger clade. We hypothesize that duplication of the C-class AGAMOUS (AG) gene followed by divergence of the duplicated AG copies during the diversification of the ginger clade lineages explains the evolution of petaloidy in the androecium. In order to address this hypothesis, we carried out phylogenetic analyses of the AG gene family across the Zingiberales and investigated patterns of gene expression within the androecium.

Results: Phylogenetic analysis supports a scenario in which Zingiberales-specific AG genes have undergone at least one round of duplication. Gene duplication was immediately followed by divergence of the retained copies. In particular, we detect positive selection in the third alpha-helix of the K domain of Zingiberales AGAMOUS copy 1 (ZinAG-1). A single fixed amino acid change is observed in ZinAG-1 within the ginger clade when compared to the banana grade. Expression analyses of AG and APETALA1/FRUITFULL (AP1/FUL) in Musa basjoo is similar to A- and C-class gene expressions in the Arabidopsis thaliana model, while Costus spicatus exhibits simultaneous expression of AG and AP1/FUL in most floral organs. We propose that this novel expression pattern could be correlated with the evolution of androecial petaloidy within the Zingiberales.

Conclusions: Our results present an intricate story in which duplication of the AG lineage has lead to the retention of at least two diverged Zingiberales-specific copies, ZinAG-1 and Zingiberales AGAMOUS copy 2 (ZinAG-2). Positive selection on ZinAG-1 residues suggests a mechanism by which AG gene divergence may explain observed morphological changes in Zingiberales flowers. Expression data provides preliminary support for the proposed mechanism, although further studies are required to fully test this hypothesis.
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http://dx.doi.org/10.1186/s13227-015-0002-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4399222PMC
April 2015

Antigen production using heterologous expression of dengue virus-2 non-structural protein 1 (NS1) in Nicotiana tabacum (Havana) for immunodiagnostic purposes.

Plant Cell Rep 2015 Jun 18;34(6):919-28. Epub 2015 Feb 18.

Laboratory of Molecular Immunovirology, Department of General Biology, Federal University of Viçosa, Av. PH Rolfs, s/n Campus Universitário, Viçosa, MG, CEP 36570-000, Brazil.

Key Message: Expression of dengue-2 virus NS1 protein in Nicotiana tabacum plants for development of dengue immunodiagnostic kits. Dengue is one of the most important diseases caused by arboviruses in the world. A significant increase in its geographical distribution has been noticed over the last 20 years, with continuous transmission of several serotypes and emergence of the hemorrhagic fever in areas where the disease was previously not prevalent. Although the methodological processes for dengue diagnosis are in deep development and improvement, a limitation for the realization of dengue diagnostic tests is the difficulty of large-scale production of the antigen to be used in diagnostic tests. Due to this demand, the purpose of this study was to obtain the non-structural protein 1 (NS1) from dengue-2 serotype by heterologous expression in Nicotiana tabacum (Havana). After confirmation of the NS1 protein gene integration in the plant genome, the heterologous protein was characterized using SDS-PAGE and immunoblotting. In an immunoenzymatic test, the recombinant NS1 protein presents an antigen potential for development of dengue immunodiagnostic kits.
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http://dx.doi.org/10.1007/s00299-015-1753-5DOI Listing
June 2015

Characterization and developmental expression of genes encoding the early carotenoid biosynthetic enzymes in Citrus paradisi Macf.

Mol Biol Rep 2012 Feb 19;39(2):895-902. Epub 2011 May 19.

Biological Sciences Department, Center for Biotechnology and Genetics, State University of Santa Cruz, Rod. Ilhéus-Itabuna km 16, Ilhéus, BA 45662-000, Brazil.

In the present study, the full-length cDNA sequences of PSY, PDS, and ZDS, encoding the early carotenoid biosynthetic enzymes in the carotenoid pathway of grapefruit (Citrus paradisi), were isolated and characterized for the first time. CpPSY contained a 1311-bp open reading frame (ORF) encoding a polypeptide of 436 amino acids, CpPDS contained a 1659-bp ORF encoding a polypeptide of 552 amino acids, and CpZDS contained a 1713-bp ORF encoding a polypeptide of 570 amino acids. Phylogenetic analysis indicated that CpPSY shares homology with PSYs from Citrus, tomato, pepper, Arabidopsis, and the monocot PSY1 group, while CpPDS and CpZDS are most closely related to orthologs from Citrus and tomato. Expression analysis revealed fluctuations in CpPSY, CpPDS, and CpZDS transcript abundance and a non-coordinated regulation between the former and the two latter genes during fruit development in albedo and juice vesicles of white ('Duncan') and red ('Flame') grapefruits. A 3× higher upregulation of CpPSY expression in juice vesicles of red-fleshed 'Flame' as compared to white-fruited 'Duncan' was observed in the middle stages of fruit development, which correlates with the well documented accumulation pattern of lycopene in red grapefruit. Together with previous data, our results suggest that the primary mechanism controlling lycopene accumulation in red grapefruit involves the transcriptional upregulation of CpPSY, which controls the flux into the carotenoid pathway, and the downregulated expression of CpLCYB2, which controls the step of cyclization of lycopene in chromoplasts during fruit ripening. A correlation between CpPSY expression and fruit color evolution in red grapefruit is demonstrated.
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http://dx.doi.org/10.1007/s11033-011-0814-2DOI Listing
February 2012

Unraveling new genes associated with seed development and metabolism in Bixa orellana L. by expressed sequence tag (EST) analysis.

Mol Biol Rep 2011 Feb 19;38(2):1329-40. Epub 2010 Jun 19.

Departamento de Ciências Biológicas, Universidade Estadual de Santa Cruz, Ilhéus, BA, Brazil.

The tropical tree Bixa orellana L. produces a range of secondary metabolites which biochemical and molecular biosynthesis basis are not well understood. In this work we have characterized a set of ESTs from a non-normalized cDNA library of B. orellana seeds to obtain information about the main developmental and metabolic processes taking place in developing seeds and their associated genes. After sequencing a set of randomly selected clones, most of the sequences were assigned with putative functions based on similarity, GO annotations and protein domains. The most abundant transcripts encoded proteins associated with cell wall (prolyl 4-hydroxylase), fatty acid (acyl carrier protein), and hormone/flavonoid (2OG-Fe oxygenase) synthesis, germination (MADS FLC-like protein) and embryo development (AP2/ERF transcription factor) regulation, photosynthesis (chlorophyll a-b binding protein), cell elongation (MAP65-1a), and stress responses (metallothionein- and thaumatin-like proteins). Enzymes were assigned to 16 different metabolic pathways related to both primary and secondary metabolisms. Characterization of two candidate genes of the bixin biosynthetic pathway, BoCCD and BoOMT, showed that they belong, respectively, to the carotenoid-cleavage dioxygenase 4 (CCD4) and caffeic acid O-methyltransferase (COMT) families, and are up-regulated during seed development. It indicates their involvement in the synthesis of this commercially important carotenoid pigment in seeds of B. orellana. Most of the genes identified here are the first representatives of their gene families in B. orellana.
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http://dx.doi.org/10.1007/s11033-010-0234-8DOI Listing
February 2011

Synergism and negative interference during co-infection of tomato and Nicotiana benthamiana with two bipartite begomoviruses.

Virology 2009 May 12;387(2):257-66. Epub 2009 Mar 12.

Departamento de Fitopatologia/BIOAGRO, Universidade Federal de Viçosa, MG, Brazil.

In Brazil, at least eight begomoviruses including Tomato rugose mosaic virus (ToRMV) and Tomato yellow spot virus (ToYSV) infect tomatoes. ToYSV symptoms in tomato and Nicotiana benthamiana appear earlier and are more severe compared to those of ToRMV. We investigated the role of several factors in this differential adaptation. To analyze infection kinetics, a single leaf was inoculated and subsequently detached after different periods of time. Viral DNA accumulation was quantified in plants, viral replication was analyzed in protoplasts, and tissue tropism was determined by in situ hybridization. Results indicate that ToYSV establishes a systemic infection and reaches a higher concentration earlier than ToRMV in both hosts. ToRMV negatively interferes with ToYSV during the initial stages of infection, but once systemic infection is established this interference ceases. In N. benthamiana, ToYSV invades the mesophyll, while ToRMV is phloem-restricted. During dual infection in this host, ToYSV releases ToRMV from the phloem.
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http://dx.doi.org/10.1016/j.virol.2009.01.046DOI Listing
May 2009

Isolation and purification of RNA from tissues rich in polyphenols, polysaccharides, and pigments of annatto (Bixa orellana L.).

Mol Biotechnol 2007 Nov 8;37(3):220-4. Epub 2007 Aug 8.

Plant Biology Department, Federal University of Viçosa, Vicosa, Minas Gerais, Brazil.

The tropical plant Bixa orellana L. (annatto) produces an array of natural products, including the pigment bixin used in the food and cosmetics industries. In order to understand the biochemical and molecular basis of the biosynthesis of these natural products, a reliable method for isolating high yields of high-quality RNA is required. Here we described a successful and reproducible method for isolation and purification of high-quantity and high-quality RNA from different tissues of annatto. This protocol overcomes the usual problems associated with large amounts of polyphenols, polysaccharides, pigments, and other secondary metabolites that are not easily removed by conventional extraction procedures. Furthermore, the proposed protocol can be easily carried out in any laboratory and it could also be extended to isolate RNA from other plant species showing similar abundance of compounds that interfere with RNA extractions. The yield and quality of the RNA were monitored by spectrophotometric analysis, separation on agarose gel, Reverse Transcription-Polymerase Chain Reaction (RT-PCR), and construction of a cDNA library.
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http://dx.doi.org/10.1007/s12033-007-0070-9DOI Listing
November 2007

Tissue-specific regulation of BiP genes: a cis-acting regulatory domain is required for BiP promoter activity in plant meristems.

Plant Mol Biol 2002 Nov;50(4-5):757-71

Departamento de Biologia Vegetal, BIOAGRO, UFV, Viçosa, MG, Brazil.

The binding protein BiP is an endoplasmic reticulum (ER)-resident member of the HSP70 stress-related protein family, which is essential for the constitutive function of the ER. In addition to responding to a variety of environmental stimuli, plant BiP exhibits a tissue-specific regulation. We have isolated two soybean BiP genomic clones, designated gsBiP6 and gsBiP9, and different extensions of their 5' flanking sequences were fused to beta-glucuronidase (GUS) reporter gene and introduced into Nicotiana tabacum by Agrobacterium tumefaciens-mediated transformation. Transgenic plants displayed prominent GUS activity in the vascular bundles of roots and shoots as well as in regions of intense cell division, such as procambial region and apical meristems. Promoter deletion analyses identified two cis-regulatory functional domains that are important for the spatially-regulated activation of BiP expression under normal plant development. While an AT-rich enhancer-like sequence, designated cis-acting regulatory domain 1, CRD1 (-358 to -211, on gsBiP6), activated expression of the BiP minimal promoter in all organs analyzed, BiP promoter activity in meristematic tissues and phloem cells required the presence of a second activating domain, CRD2 (-211 to -80). Apparently, the CRD2 sequence also harbors negative cis-acting elements, because removal of this region caused activation of gsBiP6 promoter in parenchymatic xylem rays. These results suggest that the tissue-specific control of BiP gene expression requires a complex integration of multiple cis-acting regulatory elements on the promoter.
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http://dx.doi.org/10.1023/a:1019994721545DOI Listing
November 2002
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