Silencing CAFFEOYL SHIKIMATE ESTERASE affects lignification and improves saccharification in poplar

Caffeoyl shikimate esterase (CSE) was recently shown to play an essential role in lignin biosynthesis in Arabidopsis (Arabidopsis thaliana) and later in Medicago truncatula. However, the general function of this enzyme was recently questioned by the apparent lack of CSE activity in lignifying tissues of different plant species. Here, we show that down-regulation of CSE in hybrid poplar (Populus tremula x Populus alba) resulted in up to 25% reduced lignin deposition, increased levels of p-hydroxyphenyl units in the lignin polymer, and a relatively higher cellulose content. The transgenic trees were morphologically indistinguishable from the wild type. Ultra-high-performance liquid chromatography-mass spectrometry-based phenolic profiling revealed a reduced abundance of several oligolignols containing guaiacyl and syringyl units and their corresponding hydroxycinnamaldehyde units, in agreement with the reduced flux toward coniferyl and sinapyl alcohol. These trees accumulated the CSE substrate caffeoyl shikimate along with other compounds belonging to the metabolic classes of benzenoids and hydroxycinnamates. Furthermore, the reduced lignin amount combined with the relative increase in cellulose content in the CSE down-regulated lines resulted in up to 62% more glucose released per plant upon limited saccharification when no pretreatment was applied and by up to 86% and 91% when acid and alkaline pretreatments were used. Our results show that CSE is not only important for the lignification process in poplar but is also a promising target for the development of improved lignocellulosic biomass crops for sugar platform biorefineries

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear un derstanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5–7 vast areas of the tropics remain understudied.8–11 In the American tropics, Amazonia stands out as the world’s most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepre sented in biodiversity databases.13–15 To worsen this situation, human-induced modifications16,17 may elim inate pieces of the Amazon’s biodiversity puzzle before we can use them to understand how ecological com munities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple or ganism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region’s vulnerability to environmental change. 15%–18% of the most ne glected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lostinfo:eu-repo/semantics/publishedVersio

A social and ecological assessment of tropical land uses at multiple scales: the Sustainable Amazon Network

Science has a critical role to play in guiding more sustainable development trajectories. Here, we present the Sustainable Amazon Network (Rede Amazonia Sustentavel, RAS): a multidisciplinary research initiative involving more than 30 partner organizations working to assess both social and ecological dimensions of land-use sustainability in eastern Brazilian Amazonia. The research approach adopted by RAS offers three advantages for addressing land-use sustainability problems: (i) the collection of synchronized and co-located ecological and socioeconomic data across broad gradients of past and present human use; (ii) a nested sampling design to aid comparison of ecological and socioeconomic conditions associated with different land uses across local, landscape and regional scales; and (iii) a strong engagement with a wide variety of actors and non-research institutions. Here, we elaborate on these key features, and identify the ways in which RAS can help in highlighting those problems in most urgent need of attention, and in guiding improvements in land-use sustainability in Amazonia and elsewhere in the tropics. We also discuss some of the practical lessons, limitations and realities faced during the development of the RAS initiative so far.Keywords: Social–ecological systems, Tropical forests, Land use, Interdisciplinary research, Sustainability, Trade-off

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear understanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5,6,7 vast areas of the tropics remain understudied.8,9,10,11 In the American tropics, Amazonia stands out as the world's most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepresented in biodiversity databases.13,14,15 To worsen this situation, human-induced modifications16,17 may eliminate pieces of the Amazon's biodiversity puzzle before we can use them to understand how ecological communities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple organism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region's vulnerability to environmental change. 15%–18% of the most neglected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lost

História evolutiva de um retrotransposon-LTR nos dois genomas componentes do amendoim

Dissertação (mestrado)—Universidade de Brasília, Instituto de Ciências Biológicas, Departamento de Biologia Celular, 2007.O amendoim cultivado (Arachis hypogaea) é uma cultura de grande importância econômica, sendo cultivado em vários países do mundo. As safras na agricultura são reduzidas por estresses bióticos e abióticos aos quais as espécies silvestres são resistentes. Visando a obtenção de plantas mais resistentes, foi desenvolvida uma estratégia onde as espécies silvestres seriam as doadoras de genes de resistência que seriam transferidos através de cruzamentos interespecíficos. O primeiro passo seria a construção de mapas genéticos que identifiquem as distâncias e as posições dos genes de interesse no genoma de cada espécie. O segundo passo seria a correção da diferença de ploidia entre as espécies, já que a cultivada é uma alotetraplóide (2n=4x=40) com genoma AABB e as silvestres diplóides (2n=2x=20) com genomas AA ou BB. Porém, problemas de incompatibilidade entre os cruzamentos são comuns. Isso se deve muitas vezes por causa de diferenças nas composições das regiões não-codantes e de repetições, o que faz necessário um estudo detalhado dessas regiões. Dentre as seqüências repetitivas de maior importância estão os retrotransposons, elementos que se multiplicam no genoma e inserem uma nova fita de DNA em diferentes sítios. Neste trabalho, foram isoladas e caracterizadas seqüências de um retrotransposon LTR do amendoim cultivado e de seus parentais silvestres (A. duranensis e A. ipaënsis). A seqüência codificadora da enzima transcriptase reversa foi amplificada através de PCR utilizando combinações de iniciadores específicos. O Southern blot não mostrou nenhum sinal de metilação na seqüência desse elemento, mas através de sondagem de 20 mil seqüências de ESTs nenhuma similaridade foi encontrada, sugerindo que esse elemento não é expresso. Análises filogenéticas e estatísticas foram realizadas utilizando-se 87 contigs, onde seqüências nucleotídicas e de aminoácidos formaram grupos específicos nas árvores de similaridade, indicando que o retrotransposon evoluiu diferentemente nos genomas das espécies parentais. De acordo com o cálculo do número de substituições sinônimas na seqüência desse elemento, observou-se que ele é mais variável no genoma BB. Já no genoma AA de A. duranensis ele parece ter sofrido modificações recentes, provavelmente decorridas da transposição desse elemento há cerca de 2-6 milhões de anos, fato que não pode ser observado no genoma de A. ipaënsis. O número de cópias calculado por genoma diplóide nas três espécies foi, em média, 800 em A. ipaënsis, 3 mil em A. duranensis e 5 mil em A. hypogaea. Esses resultados mostram que o retrotransposon é muito antigo e provavelmente fazia parte do genoma do ancestral comum à diferentes grupos vegetais. Após a diferenciação nas espécies parentais o retrotransposon provavelmente multiplicou seu número de cópias no genoma AA e permaneceu praticamente inalterado no genoma BB. Apesar de ser encontrado em regiões eucromáticas e de não estar metilado ele se mantém inativo. _______________________________________________________________________________ ABSTRACTThe cultivated peanut (Arachis hypogaea) is a crop of economic importance and it is cultivated in several countries around the world. Yields in agriculture are reduced by biotic and abiotic stresses to which most the wild species are resistant. With the aim of developing more resistant plants, a strategy of crossing where wild species are the donors of resistance genes to the cultivated peanut is used. The first step is to develop a linkage map to identify the distances and positions of genes of interest for each species. The second step is the correction of the number of chromosomes since the cultivated peanut is an allotetraploid (2n=4x=40) with an AABB genome and the wild species are diploid (2n=2x=20) with AA or BB genomes. However, problems of incompatibility during crosses are common. One possible cause for incompatibility is the difference in the intergenic noncoding regions of different species. For this and other reasons, the study of repetitive DNA is interesting. Among the repetitive sequences, retrotransposable elements are found to be of major importance because they can copy of themselves into new chromosome sites. In this work we reported the isolation and characterization of an LTR retrotransposon from the cultivated peanut and its wild parental genomes (A. duranensis and A. ipaënsis). The coding region of the reverse trancriptase gene was amplified by PCR using specific primer combinations derived from genomic sequences of the Arachis data base. No methylation was found using Southern blot but also no significant homology to 20.000 ESTs sequences on the GenBank, suggesting that this element is not expressed. Phylogenetic and statistical analyses were done with 87 contigs. Both the nucleotide and aminoacid sequences formed specific groups indicating that the retroelement evolved differently in the wild species genomes. According to the synonymous substitutions content it was observed that the retrotransposon has accumulated more mutations in BB genome than in the A. duranensis genome, probably because of a recent amplification that occurred about 2-6 myr ago. The copy number calculated per diploid genome for the three species was aproximately: 800 for A. ipaënsis, 3.000 for A. duranensis and 5.000 for A. hypogaea. These results show that the element is ancient and it was a part of the genome of a species ancestral to many different groups of plants. After the differentiation between the two parental species, this element probably multiplied its copy number in the AA genome and remained almost silent in the BB genome. Although the element is dispersed in euchromatic regions and does not present any methylation no sign of activity was found

Development of reference genes for RT-qPCR analysis of gene expression in Pleurotus pulmonarius for biotechnological applications

Abstract Jatropha curcas is an oilseed crop with biorefinery applications. Whilst cake generated following oil extraction offers potential as a protein source for animal feed, inactivation of toxic phorbol esters present in the material is necessary. Pleurotus pulmonarius is a detoxifying agent for jatropha cake with additional potential as animal feed, edible mushroom and for enzyme production. For the characterization of fungal genes involved in phorbol ester degradation, together with other industrial applications, reverse transcription-quantitative PCR (RT-qPCR) is a tool that enables accurate quantification of gene expression. For this, reliable analysis requires reference genes for normalization of mRNA levels validated under conditions employed for target genes. The stability of potential reference genes β-TUB, ACTIN, GAPDH, PHOS, EF1α, TRPHO, LAC, MNP3, MYP and VP were evaluated following growth of P. pulmonarius on toxic, non-toxic jatropha cake and a combined treatment, respectively. NormFinder and geNorm algorithms for expression stability analysis identified PHOS, EF1α and MNP3 as appropriate for normalizing gene expression. Reference gene combinations contrasting in ranking were compared following normalization of relative expression of the CHU_2040 gene, encoding an esterase enzyme potentially involved in phorbol ester degradation. The reference genes for P. pulmonarius will facilitate the elucidation of mechanisms involved in detoxification of phorbol esters as well as analysis of target genes for application in biorefinery models

Introducing curcumin biosynthesis in Arabidopsis enhances lignocellulosic biomass processing

Lignin is the main cause of lignocellulosic biomass recalcitrance to industrial enzymatic hydrolysis. By partially replacing the traditional lignin monomers by alternative ones, lignin extractability can be enhanced. To design a lignin that is easier to degrade under alkaline conditions, curcumin (diferuloylmethane) was produced in the model plant Arabidopsis thaliana via simultaneous expression of the turmeric (Curcuma longa) genes DIKETIDE-CoA SYNTHASE (DCS) and CURCUMIN SYNTHASE 2 (CURS2). The transgenic plants produced a plethora of curcumin- and phenylpentanoid-derived compounds with no negative impact on growth. Catalytic hydrogenolysis gave evidence that both curcumin and phenylpentanoids were incorporated into the lignifying cell wall, thereby significantly increasing saccharification efficiency after alkaline pretreatment of the transgenic lines by 14–24% as compared with the wild type. These results demonstrate that non-native monomers can be synthesized and incorporated into the lignin polymer in plants to enhance their biomass processing efficiency

Transcriptome Profiling of the Resistance Response of <i>Musa acuminata</i> subsp. <i>burmannicoides</i>, var. Calcutta 4 to <i>Pseudocercospora musae</i>

Banana (Musa spp.), which is one of the world’s most popular and most traded fruits, is highly susceptible to pests and diseases. Pseudocercospora musae, responsible for Sigatoka leaf spot disease, is a principal fungal pathogen of Musa spp., resulting in serious economic damage to cultivars in the Cavendish subgroup. The aim of this study was to characterize genetic components of the early immune response to P. musae in Musa acuminata subsp. burmannicoides, var. Calcutta 4, a resistant wild diploid. Leaf RNA samples were extracted from Calcutta 4 three days after inoculation with fungal conidiospores, with paired-end sequencing conducted in inoculated and non-inoculated controls using lllumina HiSeq 4000 technology. Following mapping to the reference M. acuminata ssp. malaccensis var. Pahang genome, differentially expressed genes (DEGs) were identified and expression representation analyzed on the basis of gene ontology enrichment, Kyoto Encyclopedia of Genes and Genomes orthology and MapMan pathway analysis. Sequence data mapped to 29,757 gene transcript models in the reference Musa genome. A total of 1073 DEGs were identified in pathogen-inoculated cDNA libraries, in comparison to non-inoculated controls, with 32% overexpressed. GO enrichment analysis revealed common assignment to terms that included chitin binding, chitinase activity, pattern binding, oxidoreductase activity and transcription factor (TF) activity. Allocation to KEGG pathways revealed DEGs associated with environmental information processing, signaling, biosynthesis of secondary metabolites, and metabolism of terpenoids and polyketides. With 144 up-regulated DEGs potentially involved in biotic stress response pathways, including genes involved in cell wall reinforcement, PTI responses, TF regulation, phytohormone signaling and secondary metabolism, data demonstrated diverse early-stage defense responses to P. musae. With increased understanding of the defense responses occurring during the incompatible interaction in resistant Calcutta 4, these data are appropriate for the development of effective disease management approaches based on genetic improvement through introgression of candidate genes in superior cultivars