Antisense Transcription in Plants: A Systematic Review and an Update on cis-NATs of Sugarcane.

Initially, natural antisense transcripts (NATs, natRNAs, or asRNAs) were considered repressors; however, their functions in gene regulation are diverse. Positive, negative, or neutral correlations to the cognate gene expression have been noted. Although the first studies were published about 50 years ago, there is still much to be investigated regarding antisense transcripts in plants. A systematic review of scientific publications available in the Web of Science databases was conducted to contextualize how the studying of antisense transcripts has been addressed. Studies were classified considering three categories: "Natural antisense" (208), artificial antisense used in "Genetic Engineering" (797), or "Natural antisense and Genetic Engineering"-related publications (96). A similar string was used for a systematic search in the NCBI Gene database. Of the 1132 antisense sequences found for plants, only 0.8% were cited in PubMed and had antisense information confirmed. This value was the lowest when compared to fungi (2.9%), bacteria (2.3%), and mice (54.1%). Finally, we present an update for the cis-NATs identified in Saccharum spp. Of the 1413 antisense transcripts found in different experiments, 25 showed concordant expressions, 22 were discordant, 1264 did not correlate with the cognate genes, and 102 presented variable results depending on the experiment.

Leaf transcriptome profiling of contrasting sugarcane genotypes for drought tolerance under field conditions.

Drought is the most detrimental abiotic stress to sugarcane production. Nevertheless, transcriptomic analyses remain scarce for field-grown plants. Here we performed comparative transcriptional profiling of two contrasting sugarcane genotypes, 'IACSP97-7065' (drought-sensitive) and 'IACSP94-2094' (drought-tolerant) grown in a drought-prone environment. Physiological parameters and expression profiles were analyzed at 42 (May) and 117 (August) days after the last rainfall. The first sampling was done under mild drought (soil water potential of -60 kPa), while the second one was under severe drought (soil water potential of -75 kPa). Microarray analysis revealed a total of 622 differentially expressed genes in both sugarcane genotypes under mild and severe drought stress, uncovering about 250 exclusive transcripts to 'IACSP94-2094' involved in oxidoreductase activity, transcriptional regulation, metabolism of amino acids, and translation. Interestingly, the enhanced antioxidant system of 'IACSP94-2094' may protect photosystem II from oxidative damage, which partially ensures stable photochemical activity even after 117 days of water shortage. Moreover, the tolerant genotype shows a more extensive set of responsive transcription factors, promoting the fine-tuning of drought-related molecular pathways. These results help elucidate the intrinsic molecular mechanisms of a drought-tolerant sugarcane genotype to cope with ever-changing environments, including prolonged water deficit, and may be useful for plant breeding programs.

Planting Season Impacts Sugarcane Stem Development, Secondary Metabolite Levels, and Natural Antisense Transcription.

To reduce the potentially irreversible environmental impacts caused by fossil fuels, the use of renewable energy sources must be increased on a global scale. One promising source of biomass and bioenergy is sugarcane. The study of this crop's development in different planting seasons can aid in successfully cultivating it in global climate change scenarios. The sugarcane variety SP80-3280 was field grown under two planting seasons with different climatic conditions. A systems biology approach was taken to study the changes on physiological, morphological, agrotechnological, transcriptomics, and metabolomics levels in the leaf +1, and immature, intermediate and mature internodes. Most of the variation found within the transcriptomics and metabolomics profiles is attributed to the differences among the distinct tissues. However, the integration of both transcriptomics and metabolomics data highlighted three main metabolic categories as the principal sources of variation across tissues: amino acid metabolism, biosynthesis of secondary metabolites, and xenobiotics biodegradation and metabolism. Differences in ripening and metabolite levels mainly in leaves and mature internodes may reflect the impact of contrasting environmental conditions on sugarcane development. In general, the same metabolites are found in mature internodes from both "one-year" and "one-and-a-half-year sugarcane", however, some metabolites (i.e., phenylpropanoids with economic value) and natural antisense transcript expression are only detected in the leaves of "one-year" sugarcane.

Amino Acid and Carbohydrate Metabolism Are Coordinated to Maintain Energetic Balance during Drought in Sugarcane.

The ability to expand crop plantations without irrigation is a major goal to increase agriculture sustainability. To achieve this end, we need to understand the mechanisms that govern plant growth responses under drought conditions. In this study, we combined physiological, transcriptomic, and genomic data to provide a comprehensive picture of drought and recovery responses in the leaves and roots of sugarcane. Transcriptomic profiling using oligoarrays and RNA-seq identified 2898 (out of 21,902) and 46,062 (out of 373,869) transcripts as differentially expressed, respectively. Co-expression analysis revealed modules enriched in photosynthesis, small molecule metabolism, alpha-amino acid metabolism, trehalose biosynthesis, serine family amino acid metabolism, and carbohydrate transport. Together, our findings reveal that carbohydrate metabolism is coordinated with the degradation of amino acids to provide carbon skeletons to the tricarboxylic acid cycle. This coordination may help to maintain energetic balance during drought stress adaptation, facilitating recovery after the stress is alleviated. Our results shed light on candidate regulatory elements and pave the way to biotechnology strategies towards the development of drought-tolerant sugarcane plants.

Assembly of the 373k gene space of the polyploid sugarcane genome reveals reservoirs of functional diversity in the world's leading biomass crop.

BACKGROUND: Sugarcane cultivars are polyploid interspecific hybrids of giant genomes, typically with 10-13 sets of chromosomes from 2 Saccharum species. The ploidy, hybridity, and size of the genome, estimated to have >10 Gb, pose a challenge for sequencing. RESULTS: Here we present a gene space assembly of SP80-3280, including 373,869 putative genes and their potential regulatory regions. The alignment of single-copy genes in diploid grasses to the putative genes indicates that we could resolve 2-6 (up to 15) putative homo(eo)logs that are 99.1% identical within their coding sequences. Dissimilarities increase in their regulatory regions, and gene promoter analysis shows differences in regulatory elements within gene families that are expressed in a species-specific manner. We exemplify these differences for sucrose synthase (SuSy) and phenylalanine ammonia-lyase (PAL), 2 gene families central to carbon partitioning. SP80-3280 has particular regulatory elements involved in sucrose synthesis not found in the ancestor Saccharum spontaneum. PAL regulatory elements are found in co-expressed genes related to fiber synthesis within gene networks defined during plant growth and maturation. Comparison with sorghum reveals predominantly bi-allelic variations in sugarcane, consistent with the formation of 2 "subgenomes" after their divergence ∼3.8-4.6 million years ago and reveals single-nucleotide variants that may underlie their differences. CONCLUSIONS: This assembly represents a large step towards a whole-genome assembly of a commercial sugarcane cultivar. It includes a rich diversity of genes and homo(eo)logous resolution for a representative fraction of the gene space, relevant to improve biomass and food production.

Novel Insights Into the Early Stages of Ratoon Stunting Disease of Sugarcane Inferred from Transcript and Protein Analysis.

Despite of the importance of ratoon stunting disease, little is known on the responses of sugarcane to its causal agent, the vascular bacterial endophyte Leifsonia xyli subsp. xyli. The transcriptome and proteome of young plants of a susceptible cultivar with no symptoms of stunting but with relative low and high bacterial titers were compared at 30 and 60 days after inoculation. Increased bacterial titers were associated with alterations in the expression of 267 cDNAs and in the abundance of 150 proteins involved in plant growth, hormone metabolism, signal transduction and defense responses. Some alterations are predicted to benefit the pathogen, such as the up-regulation of genes involved in the synthesis of methionine. Also, genes and proteins of the cell division cycle were all down-regulated in plants with higher titers at both times. It is hypothesized that the negative effects on cell division related to increased bacterial titers is cumulative over time and its modulation by other host and environmental factors results in the stunting symptom.

Building the sugarcane genome for biotechnology and identifying evolutionary trends.

BACKGROUND: Sugarcane is the source of sugar in all tropical and subtropical countries and is becoming increasingly important for bio-based fuels. However, its large (10 Gb), polyploid, complex genome has hindered genome based breeding efforts. Here we release the largest and most diverse set of sugarcane genome sequences to date, as part of an on-going initiative to provide a sugarcane genomic information resource, with the ultimate goal of producing a gold standard genome. RESULTS: Three hundred and seventeen chiefly euchromatic BACs were sequenced. A reference set of one thousand four hundred manually-annotated protein-coding genes was generated. A small RNA collection and a RNA-seq library were used to explore expression patterns and the sRNA landscape. In the sucrose and starch metabolism pathway, 16 non-redundant enzyme-encoding genes were identified. One of the sucrose pathway genes, sucrose-6-phosphate phosphohydrolase, is duplicated in sugarcane and sorghum, but not in rice and maize. A diversity analysis of the s6pp duplication region revealed haplotype-structured sequence composition. Examination of hom(e)ologous loci indicate both sequence structural and sRNA landscape variation. A synteny analysis shows that the sugarcane genome has expanded relative to the sorghum genome, largely due to the presence of transposable elements and uncharacterized intergenic and intronic sequences. CONCLUSION: This release of sugarcane genomic sequences will advance our understanding of sugarcane genetics and contribute to the development of molecular tools for breeding purposes and gene discovery.

Identification of sense and antisense transcripts regulated by drought in sugarcane.

Sugarcane is an important sugar and energy crop that can be used efficiently for biofuels production. The development of sugarcane cultivars tolerant to drought could allow for the expansion of plantations to sub-prime regions. Knowledge on the mechanisms underlying drought responses and its relationship with carbon partition would greatly help to define routes to increase yield. In this work we studied sugarcane responses to drought using a custom designed oligonucleotide array with 21,901 different probes. The oligoarrays were designed to contain probes that detect transcription in both sense and antisense orientation. We validated the results obtained using quantitative real-time PCR (qPCR). A total of 987 genes were differentially expressed in at least one sample of sugarcane plants submitted to drought for 24, 72 and 120 h. Among them, 928 were sense transcripts and 59 were antisense transcripts. Genes related to Carbohydrate Metabolism, RNA Metabolism and Signal Transduction were selected for gene expression validation by qPCR that indicated a validation percentage of 90%. From the probes presented on the array, 75% of the sense probes and 11.9% of the antisense probes have signal above background and can be classified as expressed sequences. Our custom sugarcane oligonucleotide array provides sensitivity and good coverage of sugarcane transcripts for the identification of a representative proportion of natural antisense transcripts (NATs) and sense-antisense transcript pairs (SATs). The antisense transcriptome showed, in most cases, co-expression with respective sense transcripts.