DEAD-Box RNA Helicase Family in Physic Nut (Jatropha curcas L.): Structural Characterization and Response to Salinity.

Helicases, motor proteins present in both prokaryotes and eukaryotes, play a direct role in various steps of RNA metabolism. Specifically, SF2 RNA helicases, a subset of the DEAD-box family, are essential players in plant developmental processes and responses to biotic and abiotic stresses. Despite this, information on this family in the physic nut (Jatropha curcas L.) remains limited, spanning from structural patterns to stress responses. We identified 79 genes encoding DEAD-box RNA helicases (JcDHX) in the J. curcas genome. These genes were further categorized into three subfamilies: DEAD (42 genes), DEAH (30 genes), and DExH/D (seven genes). Characterization of the encoded proteins revealed a remarkable diversity, with observed patterns in domains, motifs, and exon-intron structures suggesting that the DEAH and DExH/D subfamilies in J. curcas likely contribute to the overall versatility of the family. Three-dimensional modeling of the candidates showed characteristic hallmarks, highlighting the expected functional performance of these enzymes. The promoter regions of the JcDHX genes revealed potential cis-elements such as Dof-type, BBR-BPC, and AP2-ERF, indicating their potential involvement in the response to abiotic stresses. Analysis of RNA-Seq data from the roots of physic nut accessions exposed to 150 mM of NaCl for 3 h showed most of the JcDHX candidates repressed. The protein-protein interaction network indicated that JcDHX proteins occupy central positions, connecting events associated with RNA metabolism. Quantitative PCR analysis validated the expression of nine DEAD-box RNA helicase transcripts, showing significant associations with key components of the stress response, including RNA turnover, ribosome biogenesis, DNA repair, clathrin-mediated vesicular transport, phosphatidyl 3,5-inositol synthesis, and mitochondrial translation. Furthermore, the induced expression of one transcript (JcDHX44) was confirmed, suggesting that it is a potential candidate for future functional analyses to better understand its role in salinity stress tolerance. This study represents the first global report on the DEAD-box family of RNA helicases in physic nuts and displays structural characteristics compatible with their functions, likely serving as a critical component of the plant's response pathways.

Bridging the Gap: Combining Genomics and Transcriptomics Approaches to Understand Stylosanthes scabra, an Orphan Legume from the Brazilian Caatinga.

Stylosanthes scabra is a scientifically orphaned legume found in the Brazilian Caatinga biome (a semi-arid environment). This work utilized omics approaches to investigate some ecophysiological aspects of stress tolerance/resistance in S. scabra, study its genomic landscape, and predict potential metabolic pathways. Considering its high-confidence conceptual proteome, 1694 (~2.6%) proteins were associated with resistance proteins, some of which were found in soybean QTL regions that confer resistance to Asian soybean rust. S. scabra was also found to be a potential source of terpenes, as biosynthetic gene clusters associated with terpene biosynthesis were identified in its genome. The analysis revealed that mobile elements comprised approximately 59% of the sequenced genome. In the remaining 41% of the sections, some of the 22,681 protein-coding gene families were categorized into two informational groups: those that were specific to S. scabra and those that expanded significantly compared to their immediate ancestor. Biological process enrichment analyses indicated that these gene families play fundamental roles in the adaptation of S. scabra to extreme environments. Additionally, phylogenomic analysis indicated a close evolutionary relationship between the genera Stylosanthes and Arachis. Finally, this study found a high number (57) of aquaporin-encoding loci in the S. scabra genome. RNA-Seq and qPCR data suggested that the PIP subfamily may play a key role in the species' adaptation to water deficit conditions. Overall, these results provide valuable insights into S. scabra biology and a wealth of gene/transcript information for future legume omics studies.

Unlocking Cowpea's Defense Responses: Conserved Transcriptional Signatures in the Battle against CABMV and CPSMV Viruses.

Cowpea aphid-borne mosaic virus (CABMV) and Cowpea severe mosaic virus (CPSMV) threaten cowpea commercial production. This study aimed to analyze Conserved Transcriptional Signatures (CTS) in cowpea's genotypes that are resistant to these viruses. CTS covered up- (UR) or down-regulated (DR) cowpea transcripts in response to CABMV and CPSMV mechanical inoculations. The conservation of cowpea's UR defense response was primarily observed with the one hpi treatments, with decreased CTS representatives as time elapsed. This suggests that cowpea utilizes generic mechanisms during its early interaction with the studied viruses, and subsequently employs more specialized strategies for each viral agent. The potential action of the CTS-UR emphasizes the importance of redox balance, ethylene and jasmonic acid pathways. Additionally, the CTS-UR provides evidence for the involvement of R genes, PR proteins, and PRRs receptors-extensively investigated in combating bacterial and fungal pathogens-in the defense against viral inoculation. AP2-ERF, WRKY, and MYB transcription factors, as well as PIP aquaporins and MAPK cascades, also emerged as significant molecular players. The presented work represents the first study investigating conserved mechanisms in the cowpea defense response to viral inoculations, highlighting relevant processes for initial defense responses.

From Gene to Transcript and Peptide: A Deep Overview on Non-Specific Lipid Transfer Proteins (nsLTPs).

Non-specific lipid transfer proteins (nsLTPs) stand out among plant-specific peptide superfamilies due to their multifaceted roles in plant molecular physiology and development, including their protective functions against pathogens. These antimicrobial agents have demonstrated remarkable efficacy against bacterial and fungal pathogens. The discovery of plant-originated, cysteine-rich antimicrobial peptides such as nsLTPs has paved the way for exploring the mentioned organisms as potential biofactories for synthesizing antimicrobial compounds. Recently, nsLTPs have been the focus of a plethora of research and reviews, providing a functional overview of their potential activity. The present work compiles relevant information on nsLTP omics and evolution, and it adds meta-analysis of nsLTPs, including: (1) genome-wide mining in 12 plant genomes not studied before; (2) latest common ancestor analysis (LCA) and expansion mechanisms; (3) structural proteomics, scrutinizing nsLTPs' three-dimensional structure/physicochemical characteristics in the context of nsLTP classification; and (4) broad nsLTP spatiotemporal transcriptional analysis using soybean as a study case. Combining a critical review with original results, we aim to integrate high-quality information in a single source to clarify unexplored aspects of this important gene/peptide family.

Dehydration response in Stylosanthes scabra: Transcriptional, biochemical, and physiological modulations.

Stylosanthes scabra, popularly known as stylo, is native to the Brazilian Caatinga semiarid region and stands out as a drought-tolerant shrub forage crop. This work provides information about the plant response during the first 48 h of water deficit, followed by a rehydration treatment. Besides root transcriptomics data, 13 physiological or biochemical parameters were scrutinized. Additionally, RNA-Seq annotated transcripts not associated with the "Viridiplantae" clade were taxonomically categorized. It was found that S. scabra quickly perceives and recovers from the oscillations of the imposed water regime. Physiologically, mechanisms that minimize evapotranspiration or protect the photosynthetic apparatus stood out. Biochemically, it was found that the root tissue invests in synthesizing compounds that can act as osmolytes (proline and sugars), emphasizing the importance of osmoregulation to water deficit acclimation. Consistently, transcriptome and qPCR analyses showed that a set of enriched biological processes with upregulated (UR) transcripts were involved in protective functions against reactive oxygen species or encoding enzymes of important metabolic pathways, which might contribute to S. scabra response to water deficit. Additionally, several UR kinases and transcription factors were identified. Finally, in an innovative approach, some naturally occurring microbial groups (such as Schizosaccharomyces, Bradyrhizobium, etc.) were identified in the S. scabra roots. This study reveals insights into the physiological, biochemical, and molecular mechanisms underlying the S. scabra response to water deficit and provides candidate genes that may be useful in developing drought-tolerant crop varieties through biotechnological applications.

The WRKY transcription factor family in cowpea: Genomic characterization and transcriptomic profiling under root dehydration.

Cowpea [Vigna unguiculata (L.) Walp.] is one of the most tolerant legume crops to drought and salt stresses. WRKY transcription factor (TF) family members stand out among plant transcriptional regulators related to abiotic stress tolerance. However, little information is currently available on the expression of the cowpea WRKY gene family (VuWRKY) in response to water deficit. Thus, we analyzed genomic and transcriptomic data from cowpea to identify VuWRKY members and characterize their structure and transcriptional response under root dehydration stress. Ninety-two complete VuWRKY genes were found in the cowpea genome based on their domain characteristics. They were clustered into three groups: I (15 members), II (58), and III (16), while three genes were unclassified. Domain analysis of the encoded proteins identified four major variants of the conserved heptapeptide motif WRKYGQK. In silico analysis of VuWRKY gene promoters identified eight candidate binding motifs of cis-regulatory elements, regulated mainly by six TF families associated with abiotic stress responses. Ninety-seven VuWRKY modulated splicing variants associated with 55 VuWRKY genes were identified via RNA-Seq analysis available at the Cowpea Genomics Consortium (CpGC) database. qPCR analyses showed that 22 genes are induced under root dehydration, with VuWRKY18, 21, and 75 exhibiting the most significant induction levels. Given their central role in activating signal transduction cascades in abiotic stress response, the data provide a foundation for the targeted modification of specific VuWRKY family members to improve drought tolerance in this important climate-resilient legume in the developing world and beyond.

Lipid Transfer Proteins (LTPs)-Structure, Diversity and Roles beyond Antimicrobial Activity.

Lipid transfer proteins (LTPs) are among the most promising plant-exclusive antimicrobial peptides (AMPs). They figure among the most challenging AMPs from the point of view of their structural diversity, functions and biotechnological applications. This review presents a current picture of the LTP research, addressing not only their structural, evolutionary and further predicted functional aspects. Traditionally, LTPs have been identified by their direct isolation by biochemical techniques, whereas omics data and bioinformatics deserve special attention for their potential to bring new insights. In this context, new possible functions have been identified revealing that LTPs are actually multipurpose, with many additional predicted roles. Despite some challenges due to the toxicity and allergenicity of LTPs, a systematic review and search in patent databases, indicate promising perspectives for the biotechnological use of LTPs in human health and also plant defense.

The endophytome (plant-associated microbiome): methodological approaches, biological aspects, and biotech applications.

Similar to other organisms, plants establish interactions with a variety of microorganisms in their natural environment. The plant microbiome occupies the host plant's tissues, either internally or on its surfaces, showing interactions that can assist in its growth, development, and adaptation to face environmental stresses. The advance of metagenomics and metatranscriptomics approaches has strongly driven the study and recognition of plant microbiome impacts. Research in this regard provides comprehensive information about the taxonomic and functional aspects of microbial plant communities, contributing to a better understanding of their dynamics. Evidence of the plant microbiome's functional potential has boosted its exploitation to develop more ecological and sustainable agricultural practices that impact human health. Although microbial inoculants' development and use are promising to revolutionize crop production, interdisciplinary studies are needed to identify new candidates and promote effective practical applications. On the other hand, there are challenges in understanding and analyzing complex data generated within a plant microbiome project's scope. This review presents aspects about the complex structuring and assembly of the microbiome in the host plant's tissues, metagenomics, and metatranscriptomics approaches for its understanding, covering descriptions of recent studies concerning metagenomics to characterize the microbiome of non-model plants under different aspects. Studies involving bio-inoculants, isolated from plant microbial communities, capable of assisting in crops' productivity, are also reviewed.

The Cowpea Kinome: Genomic and Transcriptomic Analysis Under Biotic and Abiotic Stresses.

The present work represents a pioneering effort, being the first to analyze genomic and transcriptomic data from Vigna unguiculata (cowpea) kinases. We evaluated the cowpea kinome considering its genome-wide distribution and structural characteristics (at the gene and protein levels), sequence evolution, conservation among Viridiplantae species, and gene expression in three cowpea genotypes under different stress situations, including biotic (injury followed by virus inoculation-CABMV or CPSMV) and abiotic (root dehydration). The structural features of cowpea kinases (VuPKs) indicated that 1,293 bona fide VuPKs covered 20 groups and 118 different families. The RLK-Pelle was the largest group, with 908 members. Insights on the mechanisms of VuPK genomic expansion and conservation among Viridiplantae species indicated dispersed and tandem duplications as major forces for VuPKs' distribution pattern and high orthology indexes and synteny with other legume species, respectively. K a /K s ratios showed that almost all (91%) of the tandem duplication events were under purifying selection. Candidate cis-regulatory elements were associated with different transcription factors (TFs) in the promoter regions of the RLK-Pelle group. C2H2 TFs were closely associated with the promoter regions of almost all scrutinized families for the mentioned group. At the transcriptional level, it was suggested that VuPK up-regulation was stress, genotype, or tissue dependent (or a combination of them). The most prominent families in responding (up-regulation) to all the analyzed stresses were RLK-Pelle_DLSV and CAMK_CAMKL-CHK1. Concerning root dehydration, it was suggested that the up-regulated VuPKs are associated with ABA hormone signaling, auxin hormone transport, and potassium ion metabolism. Additionally, up-regulated VuPKs under root dehydration potentially assist in a critical physiological strategy of the studied cowpea genotype in this assay, with activation of defense mechanisms against biotic stress while responding to root dehydration. This study provides the foundation for further studies on the evolution and molecular function of VuPKs.

Reference genes for quantitative real-time PCR normalization of Cenostigma pyramidale roots under salt stress and mycorrhizal association.

Cenostigma pyramidale is a native legume of the Brazilian semiarid region which performs symbiotic association with arbuscular mycorrhizal fungi (AMF), being an excellent model for studying genes associated with tolerance against abiotic and biotic stresses. In RT-qPCR approach, the use of reference genes is mandatory to avoid incorrect interpretation of the relative expression. This study evaluated the stability of ten candidate reference genes (CRGs) from C. pyramidale root tissues under salt stress (three collection times) and associated with AMF (three different times of salinity). The de novo transcriptome was obtained via RNA-Seq sequencing. Three algorithms were used to calculate the stability of CRGs under different conditions: (i) global (Salt, Salt+AMF, AMF and Control, and collection times), (ii) only non-inoculated plants, and (iii) AMF (only inoculated plants). HAG2, SAC1, aRP3 were the most stable CRGs for global and AMF assays, whereas HAG2, SAC1, RHS1 were the best for salt stress assay. This CRGs were used to validate the relative expression of two up-regulated transcripts in Salt2h (RAP2-3 and PIN8). Our study provides the first set of reference genes for C. pyramidale under salinity and AMF, supporting future researches on gene expression with this species.

Transcriptome of Cenostigma pyramidale roots, a woody legume, under different salt stress times.

Salinity stress has a significant impact on the gain of plant biomass. Our study provides the first root transcriptome of Cenostigma pyramidale, a tolerant woody legume from a tropical dry forest, under three different salt stress times (30 min, 2 h, and 11 days). The transcriptome was assembled using the RNA sequencing (RNA-Seq) de novo pipeline from GenPipes. We observed 932, 804, and 3157 upregulated differentially expressed genes (DEGs) and 164, 273, and 1332 downregulated DEGs for salt over 30 min, 2 h, and 11 days, respectively. For DEGs annotated with the Viridiplantae clade in the early stress periods, the response to salt stress was mainly achieved by stabilizing homeostasis of such ions like Na+ and K+ , signaling by Ca2+ , transcription factor modulation, water transport, and oxidative stress. For salt stress at 11 days, we observed a higher modulation of transcription factors including the WRKY, MYB, bHLH, NAC, HSF, and AP2-EREBP families, as well as DEGs involved in hormonal responses, water transport, sugar metabolism, proline, and reactive oxygen scavenging mechanisms. Five selected DEGs (K+ transporter, aquaporin, glutathione S-transferase, cyclic nucleotide-gated channel, and superoxide dismutase) were validated by qPCR. Our results indicated that C. pyramidale had an early perception of salt stress modulating ionic channels and transporters, and as the stress progressed, the focus turned to the antioxidant system, aquaporins, and complex hormone responses. The results of this first root transcriptome provide clues on how this native species modulate gene expression to achieve salt stress tolerance.

Plant Thaumatin-like Proteins: Function, Evolution and Biotechnological Applications.

Thaumatin-like proteins (TLPs) are a highly complex protein family associated with host defense and developmental processes in plants, animals, and fungi. They are highly diverse in angiosperms, for which they are classified as the PR-5 (Pathogenesis-Related-5) protein family. In plants, TLPs have a variety of properties associated with their structural diversity. They are mostly associated with responses to biotic stresses, in addition to some predicted activities under drought and osmotic stresses. The present review covers aspects related to the structure, evolution, gene expression, and biotechnological potential of TLPs. The efficiency of the discovery of new TLPs is below its potential, considering the availability of omics data. Furthermore, we present an exemplary bioinformatics annotation procedure that was applied to cowpea (Vigna unguiculata) transcriptome, including libraries of two tissues (root and leaf), and two stress types (biotic/abiotic) generated using different sequencing approaches. Even without using genomic sequences, the pipeline uncovered 56 TLP candidates in both tissues and stresses. Interestingly, abiotic stress (root dehydration) was associated with a high number of modulated TLP isoforms. The nomenclature used so far for TLPs was also evaluated, considering TLP structure and possible functions identified to date. It is clear that plant TLPs are promising candidates for breeding purposes and for plant transformation aiming a better performance under biotic and abiotic stresses. The development of new therapeutic drugs against human fungal pathogens also deserves attention. Despite that, applications derived from TLP molecules are still below their potential, as it is evident in our review.

Cowpea and abiotic stresses: identification of reference genes for transcriptional profiling by qPCR.

BACKGROUND: Due to cowpea ability to fix nitrogen in poor soils and relative tolerance to drought and salt stresses, efforts have been directed to identifying genes and pathways that confer stress tolerance in this species. Real-time quantitative PCR (qPCR) has been widely used as the most reliable method to measure gene expression, due to its high accuracy and specificity. In the present study, nine candidate reference genes were rigorously tested for their application in normalization of qPCR data onto roots of four distinct cowpea accessions under two abiotic stresses: root dehydration and salt (NaCl, 100 mM). In addition, the regulation of four target transcripts, under the same referred conditions was also scrutinized. RESULTS: geNorm, NormFinder, BestKeeper, and ΔCt method results indicated a set of three statistically validated RGs for each stress condition: (I) root dehydration (actin, ubiquitin-conjugating enzyme E2 variant 1D, and a Phaseolus vulgaris unknown gene-UNK), and (II) salt (ubiquitin-conjugating enzyme E2 variant 1D, F-box protein, and UNK). The expression profile of the target transcripts suggests that flavonoids are important players in the cowpea response to the abiotic stresses analyzed, since chalcone isomerase and chalcone synthase were up-regulated in the tolerant and sensitive accessions. A lipid transfer protein also participates in the cowpea tolerance mechanisms to root dehydration and salt stress. The referred transcript was up-regulated in the two tolerant accessions and presented no differential expression in the sensitive counterparts. Chitinase B, in turn, generally related to plant defense, was an important target transcript under salt stress, being up-regulated at the tolerant, and down-regulated in the sensitive accession. CONCLUSIONS: Reference genes suitable for qPCR analyses in cowpea under root dehydration and salt stress were identified. This action will lead to a more accurate and reliable analysis of gene expression on this species. Additionally, the results obtained in this study may guide future research on gene expression in cowpea under other abiotic stress types that impose osmotic imbalance. The target genes analyzed, in turn, deserve functional evaluation due to their transcriptional regulation under stresses and biotechnological potential.

Genotype-dependent regulation of drought-responsive genes in tolerant and sensitive sugarcane cultivars.

Drought is the most damaging among the major abiotic stresses. Transcriptomic studies allow a global overview of expressed genes, providing the basis for molecular markers development. Here, the HT-SuperSAGE technique allowed the evaluation of four drought-tolerant cultivars and four-sensitive cultivars, after 24h of irrigation suppression. We identified 9831 induced unitags from roots of the tolerant cultivars with different regulations by the -sensitive cultivars after the applied stress. These unitags allowed a proposal of 15 genes, whose expressed profiles were validated by RT-qPCR, evaluating each cultivar independently. These genes covered broad metabolic processes: ethylene stress attenuation (ACCD); root growth (ß-EXP8); protein degradation [ubiquitination pathway (E2, 20SPß4); plant proteases (AP, C13)]; oxidative detoxification (TRX); fatty acid synthesis (ACC); amino acid transport (AAT), and carbohydrate metabolism [glycolysis (PFK, TPI, FBA); TCA cycle (LDP, MDH); pentose phosphate pathway (TKT)]. The expressed profiles showed a genotype-dependent regulation of the target genes. Two drought-tolerant cultivars (SP83-2847; CTC6) presented each one, nine of the induced genes. Among the -sensitive cultivars, CTC13 induced only one, while SP90-1636 induced two genes. These genes should help breeders to identify accessions managing drought stress tolerance responses, showing better ethylene stress attenuation, energy allocation, amino acid transport, and protein homeostasis.

Validation of novel reference genes for reverse transcription quantitative real-time PCR in drought-stressed sugarcane.

One of the most challenging aspects of RT-qPCR data analysis is the identification of reliable reference genes. Ideally, they should be neither induced nor repressed under different experimental conditions. To date, few reference genes have been adequately studied for sugarcane (Saccharum spp.) using statistical approaches. In this work, six candidate genes ( αTUB, GAPDH, H1, SAMDC, UBQ, and 25S rRNA) were tested for gene expression normalization of sugarcane root tissues from drought-tolerant and -sensitive accessions after continuous dehydration (24 h). By undergoing different approaches (GeNorm, NormFinder, and BestKeeper), it was shown that most of them could be used in combinations for normalization purposes, with the exception of SAMDC. Nevertheless three of them (H1, αTUB, and GAPDH) were considered the most reliable reference genes. Their suitability as reference genes validated the expression profiles of two targets (AS and PFP α1), related to SuperSAGE unitags, in agreement with results revealed by previous in silico analysis. The other two sugarcane unitags (ACC oxidase and PIP1-1), after salt stress (100 mM NaCl), presented their expressions validated in the same way. In conclusion, these reference genes will be useful for dissecting gene expression in sugarcane roots under abiotic stress, especially in transcriptomic studies using SuperSAGE or RNAseq approaches.

Early transcriptional response of soybean contrasting accessions to root dehydration.

Drought is a significant constraint to yield increase in soybean. The early perception of water deprivation is critical for recruitment of genes that promote plant tolerance. DeepSuperSAGE libraries, including one control and a bulk of six stress times imposed (from 25 to 150 min of root dehydration) for drought-tolerant and sensitive soybean accessions, allowed to identify new molecular targets for drought tolerance. The survey uncovered 120,770 unique transcripts expressed by the contrasting accessions. Of these, 57,610 aligned with known cDNA sequences, allowing the annotation of 32,373 unitags. A total of 1,127 unitags were up-regulated only in the tolerant accession, whereas 1,557 were up-regulated in both as compared to their controls. An expression profile concerning the most representative Gene Ontology (GO) categories for the tolerant accession revealed the expression "protein binding" as the most represented for "Molecular Function", whereas CDPK and CBL were the most up-regulated protein families in this category. Furthermore, particular genes expressed different isoforms according to the accession, showing the potential to operate in the distinction of physiological behaviors. Besides, heat maps comprising GO categories related to abiotic stress response and the unitags regulation observed in the expression contrasts covering tolerant and sensitive accessions, revealed the unitags potential for plant breeding. Candidate genes related to "hormone response" (LOX, ERF1b, XET), "water response" (PUB, BMY), "salt stress response" (WRKY, MYB) and "oxidative stress response" (PER) figured among the most promising molecular targets. Additionally, nine transcripts (HMGR, XET, WRKY20, RAP2-4, EREBP, NAC3, PER, GPX5 and BMY) validated by RT-qPCR (four different time points) confirmed their differential expression and pointed that already after 25 minutes a transcriptional reorganization started in response to the new condition, with important differences between both accessions.

Natural antisense transcripts in plants: a review and identification in soybean infected with Phakopsora pachyrhizi SuperSAGE library.

Natural antisense ranscripts (NAT) are RNA molecules complementary to other endogenous RNAs. They are capable of regulating the expression of target genes at different levels (transcription, mRNA stability, translation, etc.). Such a property makes them ideal for interventions in organisms' metabolism. The present study reviewed plant NAT aspects, including features, availability and genesis, conservation and distribution, coding capacity, NAT pair expression, and functions. Besides, an in silico identification of NATs pairs was presented, using deepSuperSAGE libraries of soybean infected or not with Phakopsora pachyrhizi. Results showed that around 1/3 of the 77,903 predicted trans-NATs (by PlantsNATsDB database) detected had unitags mapped in both sequences of each pair. The same 1/3 of the 436 foreseen cis-NATs showed unitags anchored in both sequences of the related pairs. For those unitags mapped in NAT pairs, a modulation expression was assigned as upregulated, downregulated, or constitutive, based on the statistical analysis (P < 0.05). As a result, the infected treatment promoted the expression of 2,313 trans-NATs pairs comprising unitags exclusively from that library (1,326 pairs had unitags only found in the mock library). To understand the regulation of these NAT pairs could be a key aspect in the ASR plant response.

New insights in the sugarcane transcriptome responding to drought stress as revealed by superSAGE.

In the scope of the present work, four SuperSAGE libraries have been generated, using bulked root tissues from four drought-tolerant accessions as compared with four bulked sensitive genotypes, aiming to generate a panel of differentially expressed stress-responsive genes. Both groups were submitted to 24 hours of water deficit stress. The SuperSAGE libraries produced 8,787,315 tags (26 bp) that, after exclusion of singlets, allowed the identification of 205,975 unitags. Most relevant BlastN matches comprised 567,420 tags, regarding 75,404 unitags with 164,860 different ESTs. To optimize the annotation efficiency, the Gene Ontology (GO) categorization was carried out for 186,191 ESTs (BlastN against Uniprot-SwissProt), permitting the categorization of 118,208 ESTs (63.5%). In an attempt to elect a group of the best tags to be validated by RTqPCR, the GO categorization of the tag-related ESTs allowed the in silico identification of 213 upregulated unitags responding basically to abiotic stresses, from which 145 presented no hits after BlastN analysis, probably concerning new genes still uncovered in previous studies. The present report analyzes the sugarcane transcriptome under drought stress, using a combination of high-throughput transcriptome profiling by SuperSAGE with the Solexa sequencing technology, allowing the identification of potential target genes during the stress response.