Introduction: Advances in Plant Omics and Systems Biology.

How the complexity of biological systems can be understood is currently limited by the amount of biological information we have available to be incorporate in the vastitude of possibilities that could represent how a biological organism function. This point of view is, of course, alive under the paradigm that describes a living thing as a whole that could never be interpreted as to the sole understanding of its separated parts.If we are going to achieve the knowledge to understand all the complex relations between the molecules, pathways, organelles, cells, organs, phenotypes, and environments is unknown. However, that is exactly what moves us toward digging the most profound nature of relationships present in the living organisms.During the last 20 years, a big workforce was dedicated to the development of techniques, instruments, and scientific approaches that guided a whole new generation of scientists into the universe of omics approaches. The implementation of technological advances in several omics applications, such as transcriptomics, proteomics, and metabolomics, has brought to light the information that nowadays reshape our previous thinking on specific aspects of plant sciences, including growth, development, organ communication, chromatin states, and metabolism, not to mention the underpinning role of regulatory mechanisms that in many cases are essentially the basis for the phenotypical expression of a biological phenomenon and plants adaptation to their environment.In this chapter, some of the original concepts of complex systems theory were briefly discussed, and examples of omics approaches that are contributing to uncovering emergent characteristics of plants are presented and discussed. The combination of several experimental and computational or mathematical approaches indicated that there is room for improvements and novel discoveries. However, the level of complexity of biological systems seems to require and demand us to unify efforts toward the integration of the large omics datasets already available and the ones to come. This unification may represent the necessary breakthrough to the achievement of the understanding of complex phenomena. To do so, the inclusion of systems biology thinking into the training of undergraduate and graduate students of plant sciences and related areas seems to be also a contribution that is necessary to be organized and implemented in a worldwide scale.

Genomic resources for energy cane breeding in the post genomics era.

Sugarcane is one of the most sustainable energy crops among cultivated crops presenting the highest tonnage of cultivated plants. Its high productivity of sugar, bioethanol and bioelectricity make it a promising green alternative to petroleum. Furthermore, the myriad of products that can be derived from sugarcane biomass has been driving breeding programs towards varieties with a higher yield of fiber and a more vigorous and sustainable performance: the energy cane. Here we provide an overview of the energy cane including plant description, breeding efforts, types, and end-uses. In addition, we describe recently published genomic resources for the development of this crop, discuss current knowledge of cell wall metabolism, bioinformatic tools and databases available for the community.

Overexpression of an evolutionarily conserved drought-responsive sugarcane gene enhances salinity and drought resilience.

BACKGROUND AND AIMS: Improving drought adaptation is more pressing for crops such as sugarcane, rice, wheat and maize, given the high dependence of these crops on irrigation. One option for enhancing adaptation to water limitation in plants is by transgenic approaches. An increasing number of genes that are associated with mechanisms used by plants to cope with water scarcity have been discovered. Genes encoding proteins with unknown functions comprise a relevant fraction of the genes that are modulated by drought. We characterized a gene in response to environmental stresses to gain insight into the unknown fraction of the sugarcane genome. Scdr2 (Sugarcane drought-responsive 2) encodes a small protein and shares highly conserved sequences within monocots, dicots, algae and fungi. METHODS: Plants overexpressing the Scdr2 sugarcane gene were examined in response to salinity and drought. Measurements of the gas exchange parameters, germination rate, water content, dry mass and oxidative damage were performed. Seeds as well as juvenile plants were used to explore the resilience level of the transgenic plants when compared with wild-type plants. KEY RESULTS: Overexpression of Scdr2 enhanced germination rates in tobacco seeds under drought and salinity conditions. Juvenile transgenic plants overexpressing Scdr2 and subjected to drought and salinity stresses showed higher photosynthesis levels, internal CO2 concentration and stomatal conductance, reduced accumulation of hydrogen peroxide in the leaves, no penalty for photosystem II and faster recovery after submission to both stress conditions. Respiration was not strongly affected by both stresses in the Scdr2 transgenic plants, whereas wild-type plants exhibited increased respiration rates. CONCLUSIONS: Scdr2 is involved in the response mechanism to abiotic stresses. Higher levels of Scdr2 enhanced resilience to salinity and drought, and this protection correlated with reduced oxidative damage. Scdr2 confers, at the physiological level, advantages to climate limitations. Therefore, Scdr2 is a potential target for improving sugarcane resilience to abiotic stress.

Ethylene-induced transcriptional and hormonal responses at the onset of sugarcane ripening.

The effects of ethephon as a sugarcane ripener are attributed to ethylene. However, the role of this phytohormone at the molecular level is unknown. We performed a transcriptome analysis combined with the evaluation of sucrose metabolism and hormone profiling of sugarcane plants sprayed with ethephon or aminoethoxyvinylglycine (AVG), an ethylene inhibitor, at the onset of ripening. The differential response between ethephon and AVG on sucrose level and sucrose synthase activity in internodes indicates ethylene as a potential regulator of sink strength. The correlation between hormone levels and transcriptional changes suggests ethylene as a trigger of multiple hormone signal cascades, with approximately 18% of differentially expressed genes involved in hormone biosynthesis, metabolism, signalling, and response. A defence response elicited in leaves favoured salicylic acid over the ethylene/jasmonic acid pathway, while the upper internode was prone to respond to ethylene with strong stimuli on ethylene biosynthesis and signalling genes. Besides, ethylene acted synergistically with abscisic acid, another ripening factor, and antagonistically with gibberellin and auxin. We identified potential ethylene target genes and characterized the hormonal status during ripening, providing insights into the action of ethylene at the site of sucrose accumulation. A molecular model of ethylene interplay with other hormones is proposed.

SNP genotyping allows an in-depth characterisation of the genome of sugarcane and other complex autopolyploids.

Many plant species of great economic value (e.g., potato, wheat, cotton, and sugarcane) are polyploids. Despite the essential roles of autopolyploid plants in human activities, our genetic understanding of these species is still poor. Recent progress in instrumentation and biochemical manipulation has led to the accumulation of an incredible amount of genomic data. In this study, we demonstrate for the first time a successful genetic analysis in a highly polyploid genome (sugarcane) by the quantitative analysis of single-nucleotide polymorphism (SNP) allelic dosage and the application of a new data analysis framework. This study provides a better understanding of autopolyploid genomic structure and is a sound basis for genetic studies. The proposed methods can be employed to analyse the genome of any autopolyploid and will permit the future development of high-quality genetic maps to assist in the assembly of reference genome sequences for polyploid species.

Effects of drought on the microtranscriptome of field-grown sugarcane plants.

Sugarcane (Saccharum spp.) is the most promising crop for renewable energy. Among the diverse stresses that affect plant productivity, drought stress frequently causes losses in sugarcane fields. Although several studies have addressed plant responses to drought using controlled environments, plant responses under field conditions are largely unknown. Recently, microRNA (miRNA)-mediated post-transcriptional regulation has been described as an important and decisive component in vegetal development and stress resistance modulation. The role of miRNAs in sugarcane responses to drought under field conditions is currently not known. Two sugarcane cultivars differing in drought tolerance were grown in the field with and without irrigation (rainfed) for 7 months. By using small RNA deep sequencing, we were able to identify 18 miRNA families comprising 30 mature miRNA sequences. Among these families, we found 13 mature miRNAs that were differentially expressed in drought-stressed plants. Seven miRNAs were differentially expressed in both cultivars. The target genes for many of the differentially expressed mature miRNAs were predicted, and some of them were validated by quantitative reverse transcription PCR. Among the targets, we found transcription factors, transporters, proteins associated with senescence, and proteins involved with flower development. All of these data increase our understanding of the role of miRNAs in the complex regulation of drought stress in field-grown sugarcane, providing valuable tools to develop new sugarcane cultivars tolerant to drought stress.

A novel stress-induced sugarcane gene confers tolerance to drought, salt and oxidative stress in transgenic tobacco plants.

BACKGROUND: Drought is a major abiotic stress that affects crop productivity worldwide. Sugarcane can withstand periods of water scarcity during the final stage of culm maturation, during which sucrose accumulation occurs. Meanwhile, prolonged periods of drought can cause severe plant losses. METHODOLOGY/PRINCIPAL FINDINGS: In a previous study, we evaluated the transcriptome of drought-stressed plants to better understand sugarcane responses to drought. Among the up-regulated genes was Scdr1 (sugarcane drought-responsive 1). The aim of the research reported here was to characterize this gene. Scdr1 encodes a putative protein containing 248 amino acids with a large number of proline (19%) and cysteine (13%) residues. Phylogenetic analysis showed that ScDR1is in a clade with homologs from other monocotyledonous plants, separate from those of dicotyledonous plants. The expression of Scdr1 in different varieties of sugarcane plants has not shown a clear association with drought tolerance. CONCLUSIONS/SIGNIFICANCE: The overexpression of Scdr1 in transgenic tobacco plants increased their tolerance to drought, salinity and oxidative stress, as demonstrated by increased photosynthesis, water content, biomass, germination rate, chlorophyll content and reduced accumulation of ROS. Physiological parameters, such as transpiration rate (E), net photosynthesis (A), stomatal conductance (gs) and internal leaf CO(2) concentration, were less affected by abiotic stresses in transgenic Scdr1 plants compared with wild-type plants. Overall, our results indicated that Scdr1 conferred tolerance to multiple abiotic stresses, highlighting the potential of this gene for biotechnological applications.

KeaA, a Dictyostelium Kelch-domain protein that regulates the response to stress and development.

BACKGROUND: The protein kinase YakA is responsible for the growth arrest and induction of developmental processes that occur upon starvation of Dictyostelium cells. yakA- cells are aggregation deficient, have a faster cell cycle and are hypersensitive to oxidative and nitrosoative stress. With the aim of isolating members of the YakA pathway, suppressors of the death induced by nitrosoative stress in the yakA- cells were identified. One of the suppressor mutations occurred in keaA, a gene identical to DG1106 and similar to Keap1 from mice and the Kelch protein from Drosophila, among others that contain Kelch domains. RESULTS: A mutation in keaA suppresses the hypersensitivity to oxidative and nitrosoative stresses but not the faster growth phenotype of yakA- cells. The growth profile of keaA deficient cells indicates that this gene is necessary for growth. keaA deficient cells are more resistant to nitrosoative and oxidative stress and keaA is necessary for the production and detection of cAMP. A morphological analysis of keaA deficient cells during multicellular development indicated that, although the mutant is not absolutely deficient in aggregation, cells do not efficiently participate in the process. Gene expression analysis using cDNA microarrays of wild-type and keaA deficient cells indicated a role for KeaA in the regulation of the cell cycle and pre-starvation responses. CONCLUSIONS: KeaA is required for cAMP signaling following stress. Our studies indicate a role for kelch proteins in the signaling that regulates the cell cycle and development in response to changes in the environmental conditions.

Sugarcane for bioenergy production: an assessment of yield and regulation of sucrose content.

An increasing number of plant scientists, including breeders, agronomists, physiologists and molecular biologists, are working towards the development of new and improved energy crops. Research is increasingly focused on how to design crops specifically for bioenergy production and increased biomass generation for biofuel purposes. The most important biofuel to date is bioethanol produced from sugars (sucrose and starch). Second generation bioethanol is also being targeted for studies to allow the use of the cell wall (lignocellulose) as a source of carbon. If a crop is to be used for bioenergy production, the crop should be high yielding, fast growing, low lignin content and requiring relatively small energy inputs for its growth and harvest. Obtaining high yields in nonprime agricultural land is a key for energy crop development to allow sustainability and avoid competition with food production. Sugarcane is the most efficient bioenergy crop of tropical and subtropical regions, and biotechnological tools for the improvement of this crop are advancing rapidly. We focus this review on the studies of sugarcane genes associated with sucrose content, biomass and cell wall metabolism and the preliminary physiological characterization of cultivars that contrast for sugar and biomass yield.

Sugarcane genes associated with sucrose content.

BACKGROUND: Sucrose content is a highly desirable trait in sugarcane as the worldwide demand for cost-effective biofuels surges. Sugarcane cultivars differ in their capacity to accumulate sucrose and breeding programs routinely perform crosses to identify genotypes able to produce more sucrose. Sucrose content in the mature internodes reach around 20% of the culms dry weight. Genotypes in the populations reflect their genetic program and may display contrasting growth, development, and physiology, all of which affect carbohydrate metabolism. Few studies have profiled gene expression related to sugarcane's sugar content. The identification of signal transduction components and transcription factors that might regulate sugar accumulation is highly desirable if we are to improve this characteristic of sugarcane plants. RESULTS: We have evaluated thirty genotypes that have different Brix (sugar) levels and identified genes differentially expressed in internodes using cDNA microarrays. These genes were compared to existing gene expression data for sugarcane plants subjected to diverse stress and hormone treatments. The comparisons revealed a strong overlap between the drought and sucrose-content datasets and a limited overlap with ABA signaling. Genes associated with sucrose content were extensively validated by qRT-PCR, which highlighted several protein kinases and transcription factors that are likely to be regulators of sucrose accumulation. The data also indicate that aquaporins, as well as lignin biosynthesis and cell wall metabolism genes, are strongly related to sucrose accumulation. Moreover, sucrose-associated genes were shown to be directly responsive to short term sucrose stimuli, confirming their role in sugar-related pathways. CONCLUSION: Gene expression analysis of sugarcane populations contrasting for sucrose content indicated a possible overlap with drought and cell wall metabolism processes and suggested signaling and transcriptional regulators to be used as molecular markers in breeding programs. Transgenic research is necessary to further clarify the role of the genes and define targets useful for sugarcane improvement programs based on transgenic plants.

Signal transduction-related responses to phytohormones and environmental challenges in sugarcane.

BACKGROUND: Sugarcane is an increasingly economically and environmentally important C4 grass, used for the production of sugar and bioethanol, a low-carbon emission fuel. Sugarcane originated from crosses of Saccharum species and is noted for its unique capacity to accumulate high amounts of sucrose in its stems. Environmental stresses limit enormously sugarcane productivity worldwide. To investigate transcriptome changes in response to environmental inputs that alter yield we used cDNA microarrays to profile expression of 1,545 genes in plants submitted to drought, phosphate starvation, herbivory and N2-fixing endophytic bacteria. We also investigated the response to phytohormones (abscisic acid and methyl jasmonate). The arrayed elements correspond mostly to genes involved in signal transduction, hormone biosynthesis, transcription factors, novel genes and genes corresponding to unknown proteins. RESULTS: Adopting an outliers searching method 179 genes with strikingly different expression levels were identified as differentially expressed in at least one of the treatments analysed. Self Organizing Maps were used to cluster the expression profiles of 695 genes that showed a highly correlated expression pattern among replicates. The expression data for 22 genes was evaluated for 36 experimental data points by quantitative RT-PCR indicating a validation rate of 80.5% using three biological experimental replicates. The SUCAST Database was created that provides public access to the data described in this work, linked to tissue expression profiling and the SUCAST gene category and sequence analysis. The SUCAST database also includes a categorization of the sugarcane kinome based on a phylogenetic grouping that included 182 undefined kinases. CONCLUSION: An extensive study on the sugarcane transcriptome was performed. Sugarcane genes responsive to phytohormones and to challenges sugarcane commonly deals with in the field were identified. Additionally, the protein kinases were annotated based on a phylogenetic approach. The experimental design and statistical analysis applied proved robust to unravel genes associated with a diverse array of conditions attributing novel functions to previously unknown or undefined genes. The data consolidated in the SUCAST database resource can guide further studies and be useful for the development of improved sugarcane varieties.

Analysis and functional annotation of an expressed sequence tag collection for tropical crop sugarcane.

To contribute to our understanding of the genome complexity of sugarcane, we undertook a large-scale expressed sequence tag (EST) program. More than 260,000 cDNA clones were partially sequenced from 26 standard cDNA libraries generated from different sugarcane tissues. After the processing of the sequences, 237,954 high-quality ESTs were identified. These ESTs were assembled into 43,141 putative transcripts. Of the assembled sequences, 35.6% presented no matches with existing sequences in public databases. A global analysis of the whole SUCEST data set indicated that 14,409 assembled sequences (33% of the total) contained at least one cDNA clone with a full-length insert. Annotation of the 43,141 assembled sequences associated almost 50% of the putative identified sugarcane genes with protein metabolism, cellular communication/signal transduction, bioenergetics, and stress responses. Inspection of the translated assembled sequences for conserved protein domains revealed 40,821 amino acid sequences with 1415 Pfam domains. Reassembling the consensus sequences of the 43,141 transcripts revealed a 22% redundancy in the first assembling. This indicated that possibly 33,620 unique genes had been identified and indicated that >90% of the sugarcane expressed genes were tagged.