Nina da Mota Soares-Cavalcanti

Expression dynamics and genome distribution of osmoprotectants in soybean: identifying important components to face abiotic stress

BackgroundDespite the importance of osmoprotectants, no previous in silico evaluation of high throughput data is available for higher plants. The present approach aimed at the identification and annotation of osmoprotectant-related sequences applied to short transcripts from a soybean HT-SuperSAGE (High Throughput Super Serial Analysis of Gene Expression; 26-bp tags) database, and also its comparison with other transcriptomic and genomic data available from different sources.MethodsA curated set of osmoprotectants related sequences was generated using text mining and selected seed sequences for identification of the respective transcripts and proteins in higher plants. To test the efficiency of the seed sequences, these were aligned against four HT-SuperSAGE contrasting libraries generated by our group using soybean tolerant and sensible plants against water deficit, considering only differentially expressed transcripts (p ≤ 0.05). Identified transcripts from soybean and their respective tags were aligned and anchored against the soybean virtual genome.ResultsThe workflow applied resulted in a set including 1,996 seed sequences that allowed the identification of 36 differentially expressed genes related to the biosynthesis of osmoprotectants [Proline (P5CS: 4, P5CR: 2), Trehalose (TPS1: 9, TPPB: 1), Glycine betaine (BADH: 4) and Myo- inositol (MIPS: 7, INPS1: 8)], also mapped in silico in the soybean genome (25 loci). Another approach considered matches using Arabidopsis full length sequences as seed sequences, and allowed the identification of 124 osmoprotectant-related sequences, matching ~10.500 tags anchored in the soybean virtual chromosomes. Osmoprotectant-related genes appeared clustered in all soybean chromosomes, with higher density in some subterminal regions and synteny among some chromosome pairs.ConclusionsSoybean presents all searched osmoprotectant categories with some important members differentially expressed among the comparisons considered (drought tolerant or sensible vs. control; tolerant vs. sensible), allowing the identification of interesting candidates for biotechnological inferences. The identified tags aligned to corresponding genes that matched 19 soybean chromosomes. Osmoprotectant-related genes are not regularly distributed in the soybean genome, but clustered in some regions near the chromosome terminals, with some redundant clusters in different chromosomes indicating their involvement in previous duplication and rearrangements events. The seed sequences, transcripts and map represent the first transversal evaluation for osmoprotectant-related genes and may be easily applied to other plants of interest.

Overall picture of expressed Heat Shock Factors in Glycine max, Lotus japonicus and Medicago truncatula

Heat shock (HS) leads to the activation of molecular mechanisms, known as HS-response, that prevent damage and enhance survival under stress. Plants have a flexible and specialized network of Heat Shock Factors (HSFs), which are transcription factors that induce the expression of heat shock proteins. The present work aimed to identify and characterize the Glycine max HSF repertory in the Soybean Genome Project (GENOSOJA platform), comparing them with other legumes (Medicago truncatula and Lotus japonicus) in view of current knowledge of Arabidopsis thaliana. The HSF characterization in leguminous plants led to the identification of 25, 19 and 21 candidate ESTs in soybean, Lotus and Medicago, respectively. A search in the SuperSAGE libraries revealed 68 tags distributed in seven HSF gene types. From the total number of obtained tags, more than 70% were related to root tissues (water deficit stress libraries vs. controls), indicating their role in abiotic stress responses, since the root is the first tissue to sense and respond to abiotic stress. Moreover, as heat stress is related to the pressure of dryness, a higher HSF expression was expected at the water deficit libraries. On the other hand, expressive HSF candidates were obtained from the library inoculated with Asian Soybean Rust, inferring crosstalk among genes associated with abiotic and biotic stresses. Evolutionary relationships among sequences were consistent with different HSF classes and subclasses. Expression profiling indicated that regulation of specific genes is associated with the stage of plant development and also with stimuli from other abiotic stresses pointing to the maintenance of HSF expression at a basal level in soybean, favoring its activation under heat-stress conditions.

In silico survey of resistance (R) genes in Eucalyptus transcriptome

A major goal of plant genome research is to recognize genes responsible for important traits. Resistance genes are among the most important gene classes for plant breeding purposes being responsible for the specific immune response including pathogen recognition, and activation of plant defence mechanisms. These genes are quite abundant in higher plants, with 210 clusters found in Eucalyptus FOREST database presenting significant homology to known R-genes. All five gene classes of R-genes with their respective conserved domains are present and expressed in Eucalyptus. Most clusters identified (93) belong to the LRR-NBS-TIR (genes with three domains: Leucine-rich-repeat, Nucleotide-binding-site and Toll interleucine 1-receptor), followed by the serine-threonine-kinase class (49 clusters). Some new combinations of domains and motifs of R-genes may be present in Eucalyptus and could represent novel gene structures. Most alignments occurred with dicots (94.3%), with emphasis on Arabidopsis thaliana (Brassicaceae) sequences. All best alignments with monocots (5.2%) occurred with rice (Oryza sativa) sequences and a single cluster aligned with the gymnosperm Pinus sylvestris (0.5%). The results are discussed and compared with available data from other crops and may bring useful evidences for the understanding of defense mechanisms in Eucalyptus and other crop species.

In silico identification of known osmotic stress responsive genes from Arabidopsis in soybean and Medicago

Plants experience various environmental stresses, but tolerance to these adverse conditions is a very complex phenomenon. The present research aimed to evaluate a set of genes involved in osmotic response, comparing soybean and medicago with the well-described Arabidopsis thaliana model plant. Based on 103 Arabidopsis proteins from 27 categories of osmotic stress response, comparative analyses against Genosoja and Medicago truncatula databases allowed the identification of 1,088 soybean and 1,210 Medicago sequences. The analysis showed a high number of sequences and high diversity, comprising genes from all categories in both organisms. Genes with unknown function were among the most representative, followed by transcription factors, ion transport proteins, water channel, plant defense, protein degradation, cellular structure, organization & biogenesis and senescence. An analysis of sequences with unknown function allowed the annotation of 174 soybean and 217 Medicago sequences, most of them concerning transcription factors. However, for about 30% of the sequences no function could be attributed using in silico procedures. The establishment of a gene set involved in osmotic stress responses in soybean and barrel medic will help to better understand the survival mechanisms for this type of stress condition in legumes.

An overall evaluation of the resistance (R) and pathogenesis-related (PR) superfamilies in soybean, as compared with Medicago and Arabidopsis

Plants have the ability to recognize and respond to a multitude of pathogens, resulting in a massive reprogramming of the plant to activate defense responses including Resistance (R) and Pathogenesis-Related (PR) genes. Abiotic stresses can also activate PR genes and enhance pathogen resistance, representing valuable genes for breeding purposes. The present work offers an overview of soybean R and PR genes present in the GENOSOJA (Brazilian Soybean Genome Consortium) platform, regarding their structure, abundance, evolution and role in the plant-pathogen metabolic pathway, as compared with Medicago and Arabidopsis. Searches revealed 3,065 R candidates (756 in Soybean, 1,142 in Medicago and 1,167 in Arabidopsis), and PR candidates matching to 1,261 sequences (310, 585 and 366 for the three species, respectively). The identified transcripts were also evaluated regarding their expression pattern in 65 libraries, showing prevalence in seeds and developing tissues. Upon consulting the SuperSAGE libraries, 1,072 R and 481 PR tags were identified in association with the different libraries. Multiple alignments were generated for Xa21 and PR-2 genes, allowing inferences about their evolution. The results revealed interesting insights regarding the variability and complexity of defense genes in soybean, as compared with Medicago and Arabidopsis.

In Silico Evaluation of Osmoprotectants in Eucalyptus Transcriptome

The synthesis of osmoprotectants is one of the main mechanisms to dribble harmful effects caused by abiotic stresses. In the present work, osmoprotectant genes were selected and used in a search for orthologs in the FORESTs database using in silico procedures. The studied genes were here analyzed for the first time in Eucalyptus , including 51 identified orthologs from nine gene families (P5CS =3, P5CR =4, TPS1 =10, TPPB =8, SAT =6, OASTL =13, CMO =1, BADH =3 and INPS1 =3). In general, the most represented genes were those involved in the trehalose and cysteine synthesis, probably due to their vital importance in the maintenance of cellular homeostasis. Overall expression pattern revealed significant amounts of reads in most libraries. Dendrograms based on analysis of selected genes from other flowering plants revealed no clear separation between mono and dicots, since diversity regarding the evolution and diversity in gene signatures may be more related to fitness and adaptation, than evolutionary patterns.

Osmoprotectants in the sugarcane (Saccharum spp.) transcriptome revealed by in silico evaluation

Environmental stresses such as drought and salinity limit crop productivity in worldwide level. These stresses often lead to the accumulation of osmoprotectants in most organisms, including plants. In the present work, a search of known osmoprotectants (P5CS, P5CR, INPS1, BADH, CMO, TPS, TPP, OASTL and SAT) was carried out in the sugarcane transcriptome (237,954 expressed sequence tags) using in silico procedures. Alignments revealed that sugarcane presents a high number of osmoprotectant candidate genes, with 56 clusters found. In silico expression revealed higher expression in stressed callus tissues and those infected by Herbaspirilum rubrisubalbicans (HR), confirming the multi-function character of the osmoprotectants. As expected, the phylogenetic analysis revealed distinct groups among angiosperms, algae, animals, fungi and bacteria, in almost all dendrograms, with high degree of sequence conservation among angiosperms. As observed in comparative analysis between the ORFs of sugarcane and other organisms, the genic structure of these plants was relatively conserved suggesting that the accumulation of compatible solutes is an ancient metabolic adaptation.

Comparative In Silico Evaluation of MYB Transcription Factors in Eucalyptus, Sugarcane and Rice Transcriptomes

Transcription factors are essential in plant growth and development regulation and during response activation to environmental stimuli. This study examined 237,954 sugarcane and 110,000 eucalyptus ESTs in search of genes related to MYB factors active during abiotic stresses in arabidopsis and rice. Searches revealed relative abundance and diversity of MYB candidate sequences in both transcriptomes. In general, 71 and 42 MYB sequences have been selected in eucalyptus and sugarcane, respectively. The generated phenogram, using selected sequences and sequences from other organisms, revealed a sub-grouping according to domain-repeats number; confirming the ancestral origin of MYB genes. Expression pattern analysis revealed higher number of reads in wood, flower and bark libraries in eucalyptus, and in tissues infected by pathogens in sugarcane. Splicing pattern comparative analyses of arabidopsis and rice ORFs with eucalyptus and sugarcane ORFs revealed that neither of them presented similarities regarding all exons and intron junctions.

Identification and expression of early nodulin in sugarcane transcriptome revealed by in silico analysis

Nodulin genes have been defined as plant genes that are induced during nodule formation in legumes. Many studies, however, revealed a number of nodulins in non-legumes, including monocot plants, suggesting that these genes play additional roles besides nodulation. The presence and expression profile of the early nodulin genes (Annexin, DMI3, NIN, NORK, CCS52A, and ENOD8) was evaluated in the sugarcane transcriptome (237,954 ESTs) using in silico procedures. 129 sugarcane clusters were identified (out of 1,476 transcripts) and their expression profile was evaluated. Higher expression was observed in libraries of flowers, roots and normalized mix of tissues, confirming their multi-function character besides the plant-bacteria endophytic interaction in sugarcane. The multiple alignments revealed high homology among sugarcane sequences and respective proteins from other plants, mainly monocots, revealing a relatively conserved genetic structure among species, probably regarding ancient genetic processes.

In silico screening for pathogenesis related-2 gene candidates in vigna unguiculata transcriptome

Plants evolved diverse mechanisms to struggle against pathogen attack, for example the activity of Pathogenesis-Related (PR) genes. Within this category PR-2 encodes a Beta-glucanase able to degrade the polysaccharides present in the pathogen cell wall. The aim of this work was to screen the NordEST database to identify PR-2 members in cowpea transcriptome and analyze the structure of the identified sequences as compared with data from public databases. After search for PR-2 sequences in NordEST; CLUSTALx and MEGA4 were used to align PR-2 orthologs and generate a dendrogram. CLUSTER program revealed the expression pattern trough differential display. A new tool was developed aiming to identify plant PR-2 proteins based in the HMMER analysis. Among results, a complete candidate from cowpea could be identified. Higher expression included all libraries submitted to biotic (cowpea severe mosaic virus, CPSMV) stress, as well as wounded and salinity stressed tissues, confirming PR expression under different kind of stresses. Dendrogram analysis showed two main clades, the outgroup and Magnoliopsida where monocots and dicots organisms were positioned as sister groups. The developed HMM model could identify PR-2 also in other important plant species, allowing the development of a bioinformatic routine that may help the identification not only of pathogenesis related genes but any other genes, classes that present similar conserved domains and motifs.