Search

The sucrose yield in sugarcane largely depends on stem morphology, including length, diameter and sugar content, making sugarcane stem a key trait in breeding. The "Bainianzhe" variety from Songxi County, Fujian Province, possesses both aerial stems and rhizomes, providing a unique model for studying stem development. We performed a spatiotemporal transcriptomic analysis of the base, middle and apical sections of both aerial stems and rhizomes. The analysis categorized transcriptomes by developmental stage-base, middle and apical-rather than environmental differences. Apical segments were enriched with genes related to cell proliferation, while base segments were linked to senescence and fibrosis. Gene regulatory networks revealed key TFs involved in stem development. Orphan genes may be involved in rhizome development through coexpression networks. Plant hormones, especially genes involved in ABA and GAs synthesis, were highly expressed in rhizomes. Thiamine-related genes were also more prevalent in rhizomes. Furthermore, the apical segments of rhizomes enriched in photosynthesis-related genes suggest adaptations to light exposure. Low average temperatures in Songxi have led to unique cold acclimation in Bainianzhe, with rhizomes showing higher expression of genes linked to unsaturated fatty acid synthesis and cold-responsive calcium signalling. This indicates that rhizomes may have enhanced cold tolerance, aiding in the plant's overwintering success., (© 2024 John Wiley & Sons Ltd.)

Sugarcane has the most complex polyploid genome in the world, and sugar-related traits are one of the most important aims in sugarcane breeding. It is essential to construct a representative pan-transcriptome that contains all transcripts of a species for studies on genetic diversity, population expression, and omics analyses in sugarcane. In this study, we constructed the first comprehensive pan-transcriptome for sugarcane, and 8434 highly reliable open reading frames were found, which were not aligned with any published sugarcane genome. The core and dispensable gene clusters, as well as high- and low-expression gene clusters of the pan-transcriptome, were identified and analyzed. The integration of two sugar content differential transcriptome data revealed nine key candidate genes, including the ScHCT gene, encoding a crucial enzyme for lignin synthesis. Furthermore, the function of the ScHCT gene was validated in Arabidopsis , which was negatively correlated with sugar content and positively correlated with lignin content. The interaction protein of ScHCT, ScABH, was screened via a yeast two-hybrid assay and further validated by point-to-point Y2H and bimolecular fluorescence complementation assays. The phenotype of the Arabidopsis abh mutant line revealed that loss of function of ABH resulted in a decrease of sucrose content in stem tissue. This study provides important reference information and genetic resources for sugarcane research and varietal improvement.

Introduction: Modern sugarcane cultivars (Saccharum spp. hybrids) derived from crosses between S. officinarum and S. spontaneum, with high-sugar traits and excellent stress tolerance inherited respectively. However, the contribution of the S. spontaneum subgenome to sucrose accumulation is still unclear., Objective: To compensate for the absence of a high-quality reference genome, a transcriptome analysis method is needed to analyze the molecular basis of differential sucrose accumulation in sugarcane hybrids and to find clues to the contribution of the S. spontaneum subgenome to sucrose accumulation., Methods: PacBio full-length sequencing was used to complement genome annotation, followed by the identification of differential genes between the high and low sugar groups using differential alternative splicing analysis and differential expression analysis. At the subgenomic level, the factors responsible for differential sucrose accumulation were investigated from the perspective of transcriptional and post-transcriptional regulation., Results: A full-length transcriptome annotated at the subgenomic level was provided, complemented by 263,378 allele-defined transcript isoforms and 139,405 alternative splicing (AS) events. Differential alternative splicing (DA) analysis and differential expression (DE) analysis identified differential genes between high and low sugar groups and explained differential sucrose accumulation factors by the KEGG pathways. In some gene models, different or even opposite expression patterns of alleles from the same gene were observed, reflecting the potential evolution of these alleles toward novel functions in polyploid sugarcane. Among DA and DE genes in the sucrose source-sink complex pathway, we found some alleles encoding sucrose accumulation-related enzymes derived from the S. spontaneum subgenome were differentially expressed or had DA events between the two contrasting sugarcane hybrids., Conclusion: Full-length transcriptomes annotated at the subgenomic level could better characterize sugarcane hybrids, and the S. spontaneum subgenome was found to contribute to sucrose accumulation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Production and hosting by Elsevier B.V.)

Drought stress can severely affect sugarcane growth and yield. The objective of this research was to identify candidate genes in sugarcane tillering seedlings in response to drought stress. We performed a comparative phenotypic, physiological and transcriptomic analysis of tiller seedlings of drought-stressed and well-watered "Guire 2" sugarcane, in a time-course experiment (5 days, 9 days and 15 days). Physiological examination reviewed that SOD, proline, soluble sugars, and soluble proteins accumulated in large amounts in tiller seedlings under different intensities of drought stress, while MDA levels remained at a stable level, indicating that the accumulation of osmoregulatory substances and the enhancement of antioxidant enzyme activities helped to limit further damage caused by drought stress. RNA-seq and weighted gene co-expression network analysis (WGCNA) were performed to identify genes and modules associated with sugarcane tillering seedlings in response to drought stress. Drought stress induced huge down-regulated in gene expression profiles, most of down-regulated genes were mainly associated with photosynthesis, sugar metabolism and fatty acid synthesis. We obtained four gene co-expression modules significantly associated with the physiological changes under drought stress (three modules positively correlated, one module negatively correlated), and found that LSG1-2, ERF1-2, SHKA, TIL, HSP18.1, HSP24.1, HSP16.1 and HSFA6A may play essential regulatory roles as hub genes in increasing SOD, Pro, soluble sugar or soluble protein contents. In addition, one module was found mostly involved in tiller stem diameter, among which members of the BHLH148 were important nodes. These results provide new insights into the mechanisms by which sugarcane tillering seedlings respond to drought stress., (© 2023. Springer Nature Limited.)

Sugarcane ( Saccharum spp. hybrid) is frequently affected by seasonal drought, which causes substantial declines in quality and yield. To understand the drought resistance mechanisms of S. officinarum , the main species of modern sugarcane, at a molecular level, we carried out a comparative analysis of transcriptome and metabolome profiling of the sugarcane variety Badila under drought stress (DS). Compared with control group (CG) plants, plants exposed to DS had 13,744 (6663 up-regulated and 7081 down-regulated) differentially expressed genes (DEGs). GO and KEGG analysis showed that the DEGs were enriched in photosynthesis-related pathways and most DEGs had down-regulated expression. Moreover, the chlorophyll content, photosynthesis (Photo), stomatal conductance (Cond), intercellular carbon dioxide concentration (Ci) and transpiration rate (Trmmol) were sharply decreased under DS. These results indicate that DS has a significant negative influence on photosynthesis in sugarcane. Metabolome analysis identified 166 (37 down-regulated and 129 up-regulated) significantly regulated metabolites (SRMs). Over 50% of SRMs were alkaloids, amino acids and their derivatives, and lipids. The five most significantly enriched KEGG pathways among SRMs were Aminoacyl-tRNA biosynthesis, 2-Oxocarboxylic acid metabolism, Biosynthesis of amino acids, Phenylalanine metabolism, and Arginine and proline metabolism ( p < 0.05). Comparing CG with DS for transcriptome and metabolome profiling (T&#95;CG/DS and M&#95;CG/DS, respectively), we found three of the same KEGG-enriched pathways, namely Biosynthesis of amino acids, Phenylalanine metabolism and Arginine and proline metabolism. The potential importance of Phenylalanine metabolism and Arginine and proline metabolism was further analyzed for response to DS in sugarcane. Seven SRMs (five up-regulated and two down-regulated) and 60 DEGs (17 up-regulated and 43 down-regulated) were enriched in Phenylalanine metabolism under DS, of which novel.31261 , Sspon.04G0008060-1A , Sspon.04G0008060-2B and Sspon.04G0008060-3C were significantly correlated with 7 SRMs. In Arginine and proline metabolism, eight SRMs (seven up-regulated and one down-regulated) and 63 DEGs (32 up-regulated and 31 down-regulated) were enriched, of which Sspon.01G0026110-1A ( OAT ) and Sspon.03G0002750-3D ( P5CS ) were strongly associated with proline (r > 0.99). These findings present the dynamic changes and possible molecular mechanisms of Phenylalanine metabolism as well as Arginine and proline metabolism under DS and provide a foundation for future research and sugarcane improvement.

Erianthus arundinaceus [Retzius] Jeswiet, a wild relative of sugarcane has a high biomass production potential and a reservoir of many genes for superior agronomic traits and tolerance to biotic and abiotic stresses. A comparative physiological, anatomical and root transcriptome analysis were carried out to identify the salt-responsive genes and metabolic pathways associated with salt-tolerant E. arundinaceus genotype IND99-907 and salinity-sensitive sugarcane genotype Co 97010. IND99-907 recorded growth of young leaves, higher proline content, higher relative water content, intact root anatomical structures and lower Na + /K + , Ca 2+ /K + and Mg 2+ /K + ratio as compared to the sugarcane genotype Co 97010. We have generated four de novo transcriptome assemblies between stressed and control root samples of IND99-907 and Co 97010. A total of 649 and 501 differentially expressed genes (FDR<0.01) were identified from the stressed and control libraries of IND99-907 and Co 97010 respectively. Genes and pathways related to early stress-responsive signal transduction, hormone signalling, cytoskeleton organization, cellular membrane stabilization, plasma membrane-bound calcium and proton transport, sodium extrusion, secondary metabolite biosynthesis, cellular transporters related to plasma membrane-bound trafficking, nucleobase transporter, clathrin-mediated endocytosis were highly enriched in IND99-907. Whereas in Co 97010, genes related to late stress-responsive signal transduction, electron transport system, senescence, protein degradation and programmed cell death, transport-related genes associated with cellular respiration and mitochondrial respiratory chain, vesicular trafficking, nitrate transporter and fewer secondary metabolite biosynthetic genes were highly enriched. A total of 27 pathways, 24 biological processes, three molecular functions and one cellular component were significantly enriched (FDR≤ 0.05) in IND99-907 as compared to 20 pathways, two biological processes without any significant molecular function and cellular components in Co 97010, indicates the unique and distinct expression pattern of genes and metabolic pathways in both genotypes. The genomic resources developed from this study is useful for sugarcane crop improvement through development of genic SSR markers and genetic engineering approaches., (© 2021. The Author(s).)

Saccharum officinarum (sugarcane) is the fifth major cultivated crop around the world. Sugarcane rind is a promising source for anthocyanin pigments; however, limited information is available on the anthocyanin and its biosynthesis in sugarcane rinds. In this study, we have quantified 49 compounds including 6 flavonoids and 43 anthocyanins in the rind of 6 sugarcane cultivars by using LCMS/MS approach. Thirty of them were quantified for the first time in sugarcane. The 43 anthocyanins included 10 cyanidin (Cya), 11 pelargonidin (Pel), 9 peonidin (Peo), 5 malvidin (Mal), 4 delphinidin (Del), and 4 petunidin (Pet) metabolites. High contents of Cya derivatives were observed in the rind of YT71/210 (dark purple rind), such as cya-3-O-(6-O-malonyl)-glu 1283.3 µg/g and cya-3-O-glu 482.67 µg/g followed by ROC22 (red rind) 821.3 µg/g and 409 µg/g, respectively, whereas the YT93/159 (green rind) showed a minimum level of these compounds. Among six cultivars, ROC22 rind has high levels of Peo derivatives such as peo-3-O-glu (197 µg/g), peo-3-O-(6-O-malonyl)-glu (69 µg/g) and peo-3-O-(6-O-p-coumaryl)-glu (55.17 µg/g). The gene expression analysis revealed that some genes, including a MYB(t) gene, were highly associated with the color phenotype. Thus, we cloned and overexpressed the gene in Arabidopsis and found the pinkish brown color in the hypocotyl of all transgenic lines compared with the wild type. Hence, we have quantified a wide range of anthocyanins in major sugarcane cultivars, reported many new anthocyanins for the first time, and concluded that Cya and Peo derivatives are the major contributing factor of dissimilar colors in sugarcane. The finding and the verification of a novel MYB gene involved in anthocyanin biosynthesis have demonstrated that our study was very valuable for gene discovery and genetic improvement of sugarcane cultivars to harvest high anthocyanin contents.

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.

Background: Although extensive breeding efforts are ongoing in sugarcane (Saccharum officinarum L.), the average yield is far below the theoretical potential. Tillering is an important component of sugarcane yield, however, the molecular mechanism underlying tiller development is still elusive. The limited genomic data in sugarcane, particularly due to its complex and large genome, has hindered in-depth molecular studies., Results: Herein, we generated full-length (FL) transcriptome from developing leaf and tiller bud samples based on PacBio Iso-Seq. In addition, we performed RNA-seq from tiller bud samples at three developmental stages (T0, T1 and T2) to uncover key genes and biological pathways involved in sugarcane tiller development. In total, 30,360 and 20,088 high-quality non-redundant isoforms were identified in leaf and tiller bud samples, respectively, representing 41,109 unique isoforms in sugarcane. Likewise, we identified 1063 and 1037 alternative splicing events identified in leaf and tiller bud samples, respectively. We predicted the presence of coding sequence for 40,343 isoforms, 98% of which was successfully annotated. Comparison with previous FL transcriptomes in sugarcane revealed 2963 unreported isoforms. In addition, we characterized 14,946 SSRs from 11,700 transcripts and 310 lncRNAs. By integrating RNA-seq with the FL transcriptome, 468 and 57 differentially expressed genes (DEG) were identified in T1vsT0 and T2vsT0, respectively. Strong up-regulation of several pyruvate phosphate dikinase and phosphoenolpyruvate carboxylase genes suggests enhanced carbon fixation and protein synthesis to facilitate tiller growth. Similarly, up-regulation of linoleate 9S-lipoxygenase and lipoxygenase genes in the linoleic acid metabolism pathway suggests high synthesis of key oxylipins involved in tiller growth and development., Conclusions: Collectively, we have enriched the genomic data available in sugarcane and provided candidate genes for manipulating tiller formation and development, towards productivity enhancement in sugarcane.

Sugarcane ratoon stunting disease (RSD) caused by Leifsonia xyli subsp. xyli (Lxx) is a common destructive disease that occurs around the world. Lxx is an obligate pathogen of sugarcane, and previous studies have reported some physiological responses of RSD-affected sugarcane. However, the molecular understanding of sugarcane response to Lxx infection remains unclear. In the present study, transcriptomes of healthy and Lxx-infected sugarcane stalks and leaves were studied to gain more insights into the gene activity in sugarcane in response to Lxx infection. RNA-Seq analysis of healthy and diseased plants transcriptomes identified 107,750 unigenes. Analysis of these unigenes showed a large number of differentially expressed genes (DEGs) occurring mostly in leaves of infected plants. Sugarcane responds to Lxx infection mainly via alteration of metabolic pathways such as photosynthesis, phytohormone biosynthesis, phytohormone action-mediated regulation, and plant-pathogen interactions. It was also found that cell wall defense pathways and protein phosphorylation/dephosphorylation pathways may play important roles in Lxx pathogeneis. In Lxx-infected plants, significant inhibition in photosynthetic processes through large number of differentially expressed genes involved in energy capture, energy metabolism and chloroplast structure. Also, Lxx infection caused down-regulation of gibberellin response through an increased activity of DELLA and down-regulation of GID1 proteins. This alteration in gibberellic acid response combined with the inhibition of photosynthetic processes may account for the majority of growth retardation occurring in RSD-affected plants. A number of genes associated with plant-pathogen interactions were also differentially expressed in Lxx-infected plants. These include those involved in secondary metabolite biosynthesis, protein phosphorylation/dephosphorylation, cell wall biosynthesis, and phagosomes, implicating an active defense response to Lxx infection. Considering the fact that RSD occurs worldwide and a significant cause of sugarcane productivity, a better understanding of Lxx resistance-related processes may help develop tools and technologies for producing RSD-resistant sugarcane varieties through conventional and/or molecular breeding., Competing Interests: No authors have competing interests.

Background: Mepiquat chloride (DPC) is a chemical that is extensively used to control internode growth and create compact canopies in cultured plants. Previous studies have suggested that DPC could also inhibit gibberellin biosynthesis in sugarcane. Unfortunately, the molecular mechanism underlying the suppressive effects of DPC on plant growth is still largely unknown., Results: In the present study, we first obtained high-quality long transcripts from the internodes of sugarcane using the PacBio Sequel System. A total of 72,671 isoforms, with N50 at 3073, were generated. These long isoforms were used as a reference for the subsequent RNA-seq. Afterwards, short reads generated from the Illumina HiSeq 4000 platform were used to compare the differentially expressed genes in both the DPC and the control groups. Transcriptome profiling showed that most significant gene changes occurred after six days post DPC treatment. These genes were related to plant hormone signal transduction and biosynthesis of several metabolites, indicating that DPC affected multiple pathways, in addition to suppressing gibberellin biosynthesis. The network of DPC on the key stage was illustrated by weighted gene co-expression network analysis (WGCNA). Among the 36 constructed modules, the top positive correlated module, at the stage of six days post spraying DPC, was sienna3. Notably, Stf0 sulfotransferase, cyclin-like F-box, and HOX12 were the hub genes in sienna3 that had high correlation with other genes in this module. Furthermore, the qPCR validated the high accuracy of the RNA-seq results., Conclusion: Taken together, we have demonstrated the key role of these genes in DPC-induced growth inhibition in sugarcane.

Background: The development of biomass crops aims to meet industrial yield demands, in order to optimize profitability and sustainability. Achieving these goals in an energy crop like sugarcane relies on breeding for sucrose accumulation, fiber content and stalk number. To expand the understanding of the biological pathways related to these traits, we evaluated gene expression of two groups of genotypes contrasting in biomass composition., Results: First visible dewlap leaves were collected from 12 genotypes, six per group, to perform RNA-Seq. We found a high number of differentially expressed genes, showing how hybridization in a complex polyploid system caused extensive modifications in genome functioning. We found evidence that differences in transposition and defense related genes may arise due to the complex nature of the polyploid Saccharum genomes. Genotypes within both biomass groups showed substantial variability in genes involved in photosynthesis. However, most genes coding for photosystem components or those coding for phosphoenolpyruvate carboxylases (PEPCs) were upregulated in the high biomass group. Sucrose synthase (SuSy) coding genes were upregulated in the low biomass group, showing that this enzyme class can be involved with sucrose synthesis in leaves, similarly to sucrose phosphate synthase (SPS) and sucrose phosphate phosphatase (SPP). Genes in pathways related to biosynthesis of cell wall components and expansins coding genes showed low average expression levels and were mostly upregulated in the high biomass group., Conclusions: Together, these results show differences in carbohydrate synthesis and carbon partitioning in the source tissue of distinct phenotypic groups. Our data from sugarcane leaves revealed how hybridization in a complex polyploid system resulted in noticeably different transcriptomic profiles between contrasting genotypes.

WRKY is one of the largest transcription factor families in plants and plays important roles in the regulation of developmental and physiological processes. To date, the WRKY gene family has not been identified in Saccharum species because of its complex polyploid genome. In this study, a total of 294 sequences for 154 SsWRKY genes were identified in the polyploid Saccharum spontaneum genome and then named on the basis of their chromosome locations, including 13 (8.4%) genes with four alleles, 29 (18.8%) genes with three alleles and 41 (26.6%) genes with two alleles. Among them, 73.8% and 16.0% of the SsWRKY genes originated from segmental duplications and tandem duplications, respectively. The WRKY members exhibited conserved gene structures and amino acid sequences among the allelic haplotypes, which were accompanied by variations in intron sizes. Phylogenetic and collinearity analyses revealed that 27 SsWRKYs originated after the split of sorghum and Saccharum, resulting in a significantly higher number of WRKYs in sugarcane than in the proximal diploid species sorghum. The analysis of RNA-seq data revealed that SsWRKYs' expression profiles in 46 different samples including different developmental stages revealed distinct temporal and spatial patterns with 52 genes expressed in all tissues, four genes not expressed in any tissues and 21 SsWRKY genes likely to be involved in photosynthesis. The comprehensive analysis of SsWRKYs' expression will provide an important and valuable foundation for further investigation of the regulatory mechanisms of WRKYs in physiological roles in sugarcane S. spontaneum., (The Author(s) 2019. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Sugarcane ( Saccharum spp. hybrids) is a major source of sugar and renewable bioenergy crop worldwide and suffers serious yield losses due to many pathogen infections. Leaf scald caused by Xanthomonas albilineans is a major bacterial disease of sugarcane in most sugarcane-planting countries. The molecular mechanisms of resistance to leaf scald in this plant are, however, still unclear. We performed a comparative transcriptome analysis between resistant (LCP 85-384) and susceptible (ROC20) sugarcane cultivars infected by X. albilineans using the RNA-seq platform. 24 cDNA libraries were generated with RNA isolated at four time points (0, 24, 48, and 72 h post inoculation) from the two cultivars with three biological replicates. A total of 105,783 differentially expressed genes (DEGs) were identified in both cultivars and the most upregulated and downregulated DEGs were annotated for the processes of the metabolic and single-organism categories, respectively. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the 7612 DEGs showed that plant-pathogen interaction, spliceosome, glutathione metabolism, protein processing in endoplasmic reticulum, and plant hormone signal transduction contributed to sugarcane's response to X. albilineans infection. Subsequently, relative expression levels of ten DEGs determined by quantitative reverse transcription-PCR (qRT-PCR), in addition to RNA-Seq data, indicated that different plant hormone (auxin and ethylene) signal transduction pathways play essential roles in sugarcane infected by X. albilineans. In conclusion, our results provide, for the first time, valuable information regarding the transcriptome changes in sugarcane in response to infection by X. albilineans , which contribute to the understanding of the molecular mechanisms underlying the interactions between sugarcane and this pathogen and provide important clues for further characterization of leaf scald resistance in sugarcane.

Although several metallic elements are required for plant growth, excessive amounts of aluminum ions (Al3+) can result in the inhibition of root growth, thus triggering water and nutrient deficiencies. Plants under stress undergo gene expression changes in specific genes or post-transcriptional gene regulators, such as miRNAs, that can lead to stress tolerance. In this study, we investigated the miRNAs involved in the response of sugarcane to aluminum stress. Four miRNA libraries were generated using sugarcane roots of one tolerant and one sensitive sugarcane cultivar grown under aluminum stress and used to identify the miRNAs involved in the sugarcane aluminum toxicity response. The contrast in field phenotypes of sugarcane cultivars in the field during aluminum stress was reflected in the micro-transcriptome expression profiles. We identified 394 differentially expressed miRNAs in both cultivars, 104 of which were tolerant cultivar-specific, 116 were sensitive cultivar-specific, and 87 of which were common among cultivars. In addition, 52% of differentially expressed miRNAs were upregulated in the tolerant cultivar while the majority of differentially expressed miRNAs in the sensitive cultivar were downregulated. Real-time quantitative polymerase chain reaction was used to validate the expression levels of differentially expressed miRNAs. We also attempted to identify target genes of miRNAs of interest. Our results show that selected differentially expressed miRNAs of aluminum-stressed sugarcane cultivars play roles in signaling, root development, and lateral root formation. These genes thus may be important for aluminum tolerance in sugarcane and could be used in breeding programs to develop tolerant cultivars., Competing Interests: The authors have declared that no competing interests exist.

Sugarcane is an important sugar and energy crop worldwide. It utilises highly efficient C4 photosynthesis and accumulates sucrose in its culms. The sucrose content in sugarcane culms is a quantitative trait controlled by multiple genes. The regulatory mechanism underlying the maximum sucrose level in sugarcane culms remains unclear. We used transcriptome sequences to identify the potential regulatory genes involved in sucrose accumulation in Saccarum officinarum L. cv. Badila. The sucrose accumulating internodes at the elongation and mature growth stage and the immature internodes with low sucrose content at the mature stage were used for RNA sequencing. The obtained differentially expressed genes (DEGs) related to sucrose accumulation were analysed. Results showed that the transcripts encoding invertase (beta-fructofuranosidase, EC: 3.2.1.26) which catalyses sucrose hydrolysis and 6-phosphofructokinase (PFK, EC: 2.7.1.11), a key glycolysis regulatory enzyme, were downregulated in the high sucrose accumulation internodes. The transcripts encoding key enzymes for ABA, gibberellin and ethylene synthesis were also downregulated during sucrose accumulation. Furthermore, regulated protein kinase, transcription factor and sugar transporter genes were also obtained. This research can clarify the molecular regulation network of sucrose accumulation in sugarcane., (Copyright © 2019 Elsevier Masson SAS. All rights reserved.)

Background: Sugarcane accumulates very high levels of sucrose in the culm. Elucidation of the molecular mechanisms that allows such high sucrose synthesis and accumulation (up to 650 mM) is made difficult by the complexity of the highly polyploid genome. Here we report the use of RNA Seq data to characterize the sucrose synthase (SuSy) genes expressed in the transcriptome of the mature sugarcane plant., Results: Four SuSy gene families were identified in the sugarcane Iso-Seq long read transcriptome (SUGIT) through gene annotation of transcripts that mapped to reference SuSy genes from sorghum and maize. In total, 38, 19, 14, and 2 transcripts were identified for the four corresponding SuSy genes 1, 2, 4 and 7, respectively. Comparative studies using available SuSy genes from sorghum (1, 2, 4, 6, 7) and maize (1-7) revealed that the sugarcane SuSy genes were interrupted by multiple introns and that they share a highly conserved gene structure. Spatial expression of the four SuSy genes in sugarcane genotypes and in the progenitor species, Saccharum spontaneum and Saccharum officinarum, was studied in the leaf and root tissues and also in three regions of the culm tissue; top, middle and bottom internodes. Expression profiles indicated that all SuSy transcripts were differentially expressed between the top and bottom tissues, with high expression in the top tissues, lower expression in the bottom and moderate expression in the middle, indicating a gradient of SuSy activity in the sugarcane culm. Further, the root tissue had similar expression levels to that of the top internodes while leaf tissues showed lower expression. In the progenitors, SuSy7 was found to be highly expressed in S. officinarum while the other three SuSy genes had moderate expression in both the progenitors., Conclusions: The high expression of the SuSy genes in sink tissues, the top internodes and the roots suggests functional roles in sucrose utilization to support growth. The SuSy7 gene has not been previously reported in sugarcane. As sugarcane is unique in storing such high amounts of sucrose, it is possible that there are more SuSy genes/isoforms with specific expression patterns to be discovered in this complex system.

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.

Background: Sugarcane is an important sugar and energy crop that is widely planted in the world. Among the environmental stresses, the water-deficit stress is the most limiting to plant productivity. Some groups have used PCR-based and microarray technologies to investigate the gene expression changes of multiple sugarcane cultivars under water stress. Our knowledge about sugarcane genes in response to water deficit is still poor., Results: A wild sugarcane type, Saccharum narenga, was selected and treated with drought stress for 22 days. Leaves from drought treated (DTS) and control (CK) plants were obtained for deep sequencing. Paired-end sequencing enabled us to assemble 104,644 genes (N50 = 1605 bp), of which 38,721 were aligned to other databases, such as UniProt, NR, GO, KEGG and Pfam. Single-end and paired-end sequencing identified 30,297 genes (> 5 TPM) in all samples. Compared to CK, 3389 differentially expressed genes (DEGs) were identified in DTS samples, comprising 1772 up-regulated and 1617 down-regulated genes. Functional analysis showed that the DEGs were involved in biological pathways like response to blue light, metabolic pathways and plant hormone signal transduction. We further observed the expression patterns of several important gene families, including aquaporins, late embryogenesis abundant proteins, auxin related proteins, transcription factors (TFs), heat shock proteins, light harvesting chlorophyll a-b binding proteins, disease resistance proteins, and ribosomal proteins. Interestingly, the regulation of genes varied among different subfamilies of aquaporin and ribosomal proteins. In addition, DIVARICATA and heat stress TFs were first reported in sugarcane leaves in response to water deficit. Further, we showed potential miRNAs that might be involved in the regulation of gene changes in sugarcane leaves under the water-deficit stress., Conclusions: This is the first transcriptome study of Saccharum narenga and the assembled genes are a valuable resource for future research. Our findings will improve the understanding of the mechanism of gene regulation in sugarcane leaves under the water-deficit stress. The output of this study will also contribute to the sugarcane breeding program.

Sugarcane is among the most efficient crops in converting solar energy into chemical energy. However, due to its complex genome structure and inheritance, the genetic and molecular basis of biomass yield in sugarcane is still largely unknown. We created an F2 segregating population by crossing S. officinarum and S. spontaneum and evaluated the biomass yield of the F2 individuals. The F2 individuals exhibited clear transgressive segregation in biomass yield. We sequenced transcriptomes of source and sink tissues from 12 selected extreme segregants to explore the molecular basis of high biomass yield for future breeding of high-yielding energy canes. Among the 103,664 assembled unigenes, 10,115 and 728 showed significant differential expression patterns between the two extreme segregating groups in the top visible dewlap leaf and the 9 th culm internode, respectively. The most enriched functional categories were photosynthesis and fermentation in the high-biomass and the low-biomass groups, respectively. Our results revealed that high-biomass yield was mainly determined by assimilation of carbon in source tissues. The high-level expression of fermentative genes in the low-biomass group was likely induced by their low-energy status. Group-specific expression alleles which can be applied in the development of new high-yielding energy cane varieties via molecular breeding were identified.

Background: Sugarcane is a major crop of the tropics cultivated mainly for its high sucrose content. The crop is genetically less explored due to its complex polyploid genome. Sucrose synthesis and accumulation are complex processes influenced by physiological, biochemical and genetic factors, and the growth environment. The recent focus on the crop for fibre and biofuel has led to a renewed interest on understanding the molecular basis of sucrose and biomass traits. This transcriptome study aimed to identify genes that are associated with and differentially regulated during sucrose synthesis and accumulation in the mature stage of sugarcane. Patterns of gene expression in high and low sugar genotypes as well as mature and immature culm tissues were studied using RNA-Seq of culm transcriptomes., Results: In this study, 28 RNA-Seq libraries from 14 genotypes of sugarcane differing in their sucrose content were used for studying the transcriptional basis of sucrose accumulation. Differential gene expression studies were performed using SoGI (Saccharum officinarum Gene Index, 3.0), SAS (sugarcane assembled sequences) of sugarcane EST database (SUCEST) and SUGIT, a sugarcane Iso-Seq transcriptome database. In total, about 34,476 genes were found to be differentially expressed between high and low sugar genotypes with the SoGI database, 20,487 genes with the SAS database and 18,543 genes with the SUGIT database at FDR < 0.01, using the Baggerley's test. Further, differential gene expression analyses were conducted between immature (top) and mature (bottom) tissues of the culm. The DEGs were functionally annotated using GO classification and the genes consistently associated with sucrose accumulation were identified., Conclusions: The large number of DEGs may be due to the large number of genes that influence sucrose content or are regulated by sucrose content. These results indicate that apart from being a primary metabolite and storage and transport sugar, sucrose may serve as a signalling molecule that regulates many aspects of growth and development in sugarcane. Further studies are needed to confirm if sucrose regulates the expression of the identified DEGs or vice versa. The DEGs identified in this study may lead to identification of genes/pathways regulating sucrose accumulation and/or regulated by sucrose levels in sugarcane. We propose identifying the master regulators of sucrose if any in the future.

Sugarcane production is strongly influenced by drought, which is a limiting factor for agricultural productivity in the world. In this study, the gene expression profiles obtained by de novo assembly of the leaf transcriptome of two sugarcane cultivars that differ in their physiological response to water deficit were evaluated by the RNA-Seq method: drought-tolerant cultivar (SP81-3250) and drought-sensitive cultivar (RB855453). For this purpose, plants were grown in a greenhouse for 60 days and were then submitted to three treatments: control (-0.01 to -0.015 MPa), moderate water deficit (-0.05 to -0.055 MPa), and severe water deficit (-0.075 to -0.08 MPa). The plants were evaluated 30, 60, and 90 days after the beginning of treatment. Sequencing on an Illumina platform (RNA-Seq) generated more than one billion sequences, resulting in 177,509 and 185,153 transcripts for the tolerant and sensitive cultivar, respectively. These transcripts were aligned with sequences from Saccharum spp, Sorghum bicolor, Miscanthus giganteus, and Arabidopsis thaliana available in public databases. The differentially expressed genes detected during the prolonged period of water deficit permit to increase our understanding of the molecular patterns involved in the physiological response of the two cultivars. The tolerant cultivar differentially expressed a larger number of genes at 90 days, while in the sensitive cultivar the number of differentially expressed genes was higher in 30 days. Both cultivars perceived the lack of water, but the tolerant cultivar responded more slowly than the sensitive cultivar. The latter requires rapid activation of different water-deficit stress response mechanisms for its survival. This rapid activation of metabolic pathways in response to water stress does not appear to be the key mechanism of drought tolerance in sugarcane. There is still much to clarify on the molecular and physiological pattern of plants in response to drought.

Sugarcane is an important tropical crop mainly cultivated to produce ethanol and sugar. Crop productivity is negatively affected by Acidovorax avenae subsp avenae (Aaa), which causes the red stripe disease. Little is known about the molecular mechanisms triggered in response to the infection. We have investigated the molecular mechanism activated in sugarcane using a RNA-seq approach. We have produced a de novo transcriptome assembly (TR7) from sugarcane RNA-seq libraries submitted to drought and infection with Aaa. Together, these libraries present 247 million of raw reads and resulted in 168,767 reference transcripts. Mapping in TR7 of reads obtained from infected libraries, revealed 798 differentially expressed transcripts, of which 723 were annotated, corresponding to 467 genes. GO and KEGG enrichment analysis showed that several metabolic pathways, such as code for proteins response to stress, metabolism of carbohydrates, processes of transcription and translation of proteins, amino acid metabolism and biosynthesis of secondary metabolites were significantly regulated in sugarcane. Differential analysis revealed that genes in the biosynthetic pathways of ET and JA PRRs, oxidative burst genes, NBS-LRR genes, cell wall fortification genes, SAR induced genes and pathogenesis-related genes (PR) were upregulated. In addition, 20 genes were validated by RT-qPCR. Together, these data contribute to a better understanding of the molecular mechanisms triggered by the Aaa in sugarcane and opens the opportunity for the development of molecular markers associated with disease tolerance in breeding programs., Competing Interests: FASTERIS SA provided support in the form of salaries for authors [L.F.], but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The contribution of L.F. (from FASTERIS SA) does not alter our adherence to PLOS ONE policies on sharing data and materials.

Saccharum spontaneum L., a wild relative of sugarcane, is known for its adaptability to environmental stresses, particularly cold stress. In the present study, an attempt was made for transcriptome profiling of the low temperature (10°C) tolerant S. spontaneum clone IND 00-1037 collected from high altitude regions of Arunachal Pradesh, North Eastern India. The Illumina Nextseq500 platform yielded a total of 47.63 and 48.18 million reads corresponding to 4.7 and 4.8 gigabase pairs (Gb) of processed reads for control and cold stressed (10°C for 24h) samples, respectively. These reads were de novo assembled into 214,611 unigenes with an average length of 801bp. Further, all unigenes were aligned to GO, KEGG and COG databases in order to identify novel genes and pathways responsive upon low temperature conditions. The differential gene expression analysis revealed that about 2583 genes were upregulated and 3302 genes were down regulated during the stress. This is perhaps the comprehensive transcriptome data of a low temperature tolerant clone of S. spontaneum. This study would aid in identifying novel genes and also in future genomic studies pertaining to sugarcane and its wild relatives., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: Sugarcane (Saccharum officinarum L.) is an important sugar crop which belongs to the grass family and can be used for fuel ethanol production. The growing demands for sugar and biofuel is asking for breeding a sugarcane variety that can shed their leaves during the maturity time due to the increasing cost on sugarcane harvest., Results: To determine leaf abscission related genes in sugarcane, we generated 524,328,950 paired reads with RNA-Seq and profiled the transcriptome of new born leaves of leaf abscission sugarcane varieties (Q1 and T) and leaf packaging sugarcane varieties (Q2 and B). Initially, 275,018 transcripts were assembled with N50 of 1,177 bp. Next, the transcriptome was annotated by mapping them to NR, UniProtKB/Swiss-Prot, Gene Ontology and KEGG pathway databases. Further, we used TransDecoder and Trinotate to obtain the likely proteins and annotate them in terms of known proteins, protein domains, signal peptides, transmembrane regions and rRNA transcripts. Different expression analysis showed 1,202 transcripts were up regulated in leaf abscission sugarcane varieties, relatively to the leaf packaging sugarcane varieties. Functional analysis told us 62, 38 and 10 upregulated transcripts were involved in plant-pathogen interaction, response to stress and abscisic acid associated pathways, respectively. The upregulation of transcripts encoding 4 disease resistance proteins (RPM1, RPP13, RGA2, and RGA4), 6 ABC transporter G family members and 16 transcription factors including WRK33 and heat stress transcription factors indicate they may be used as candidate genes for sugarcane breeding. The expression levels of transcripts were validated by qRT-PCR. In addition, we characterized 3,722 SNPs between leaf abscission and leaf packaging sugarcane plants., Conclusion: Our results showed leaf abscission associated genes in sugarcane during the maturity period. The output of this study provides a valuable resource for future genetic and genomic studies in sugarcane.

Sugarcane is an important crop worldwide for sugar and first generation ethanol production. Recently, the residue of sugarcane mills, named bagasse, has been considered a promising lignocellulosic biomass to produce the second-generation ethanol. Lignin is a major factor limiting the use of bagasse and other plant lignocellulosic materials to produce second-generation ethanol. Lignin biosynthesis pathway is a complex network and changes in the expression of genes of this pathway have in general led to diverse and undesirable impacts on plant structure and physiology. Despite its economic importance, sugarcane genome was still not sequenced. In this study a high-throughput transcriptome evaluation of two sugarcane genotypes contrasting for lignin content was carried out. We generated a set of 85,151 transcripts of sugarcane using RNA-seq and de novo assembling. More than 2,000 transcripts showed differential expression between the genotypes, including several genes involved in the lignin biosynthetic pathway. This information can give valuable knowledge on the lignin biosynthesis and its interactions with other metabolic pathways in the complex sugarcane genome.

Transcriptome analysis of sugarcane hybrid CP72-1210 (cold susceptible) and Saccharum spontaneum TUS05-05 (cold tolerant) using Sugarcane Assembled Sequences (SAS) from SUCEST-FUN Database showed that a total of 35,340 and 34,698 SAS genes, respectively, were expressed before and after chilling stress. The analysis revealed that more than 600 genes are differentially expressed in each genotype after chilling stress. Blast2Go annotation revealed that the major difference in gene expression profiles between CP72-1210 and TUS05-05 after chilling stress are present in the genes related to the transmembrane transporter activity. To further investigate the relevance of transmembrane transporter activity against abiotic stress tolerance, a S. spontaneum homolog of a NOD26-like major intrinsic protein gene (SspNIP2) was selected for functional analysis, of which expression was induced after chilling stress in the cold tolerant TUS05-05. Quantitative real-time PCR showed that SspNIP2 expression was increased ~2.5 fold at 30 minutes after cold treatment and stayed induced throughout the 24 hours of cold treatment. The amino acid sequence analysis of the cloned SspNIP2 confirmed the presence of six transmembrane domains and two NPA (Asn-Pro-Ala) motifs, signature features of major intrinsic protein families. Amino acid analysis confirmed that four amino acids, comprising the ar/R (aromatic residue/arginine) region responsible for the substrate specificity among MIPs, are conserved among monocot silicon transporters and SspNIP2. Salinity stress test on SspNIP2 transgenic tobacco plants resulted in more vigorous transgenic lines than the non-transgenic tobacco plants, suggesting some degree of tolerance to salt stress conferred by SspNIP2. SspNIP2-transgenic plants, exposed to 2 weeks of water stress without irrigation, developed various degrees of water stress symptom. The water stress test confirmed that the SspNIP2 transgenic lines had lower evapotranspiration rates than non-transgenic lines, suggesting that SspNIP2 transgenic lines showed a slight tolerance to the early water stress compared to wild type plants.

Background: Miscanthus is a promising biomass crop for temperate regions. Despite the increasing interest in this plant, limited sequence information has constrained research into its biology, physiology, and breeding. The whole genome transcriptomes of M. sinensis and M. sacchariflorus presented in this study may provide good resources to understand functional compositions of two important Miscanthus genomes and their evolutionary relationships., Results: For M. sinensis, a total of 457,891 and 512,950 expressed sequence tags (ESTs) were produced from leaf and rhizome tissues, respectively, which were assembled into 12,166 contigs and 89,648 singletons for leaf, and 13,170 contigs and 112,138 singletons for rhizome. For M. sacchariflorus, a total of 288,806 and 267,952 ESTs from leaf and rhizome tissues, respectively, were assembled into 8,732 contigs and 66,881 singletons for leaf, and 8,104 contigs and 63,212 singletons for rhizome. Based on the distributions of synonymous nucleotide substitution (Ks), sorghum and Miscanthus diverged about 6.2 million years ago (MYA), Saccharum and Miscanthus diverged 4.6 MYA, and M. sinensis and M. sacchariflorus diverged 1.5 MYA. The pairwise alignment of predicted protein sequences from sorghum-Miscanthus and two Miscanthus species found a total of 43,770 and 35,818 nsSNPs, respectively. The impacts of striking mutations found by nsSNPs were much lower between sorghum and Miscanthus than those between the two Miscanthus species, perhaps as a consequence of the much higher level of gene duplication in Miscanthus and resulting ability to buffer essential functions against disturbance., Conclusions: The ESTs generated in the present study represent a significant addition to Miscanthus functional genomics resources, permitting us to discover some candidate genes associated with enhanced biomass production. Ks distributions based on orthologous ESTs may serve as a guideline for future research into the evolution of Miscanthus species as well as its close relatives sorghum and Saccharum.

Sugarcane is an important crop and a major source of sugar and alcohol. In this study, we performed de novo assembly and transcriptome annotation for six sugarcane genotypes involved in bi-parental crosses. The de novo assembly of the sugarcane transcriptome was performed using short reads generated using the Illumina RNA-Seq platform. We produced more than 400 million reads, which were assembled into 72,269 unigenes. Based on a similarity search, the unigenes showed significant similarity to more than 28,788 sorghum proteins, including a set of 5,272 unigenes that are not present in the public sugarcane EST databases; many of these unigenes are likely putative undescribed sugarcane genes. From this collection of unigenes, a large number of molecular markers were identified, including 5,106 simple sequence repeats (SSRs) and 708,125 single-nucleotide polymorphisms (SNPs). This new dataset will be a useful resource for future genetic and genomic studies in this species.

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.

Salt stress is a primary cause of crop losses worldwide, and it has been the subject of intense investigation to unravel the complex mechanisms responsible for salinity tolerance. MicroRNA is implicated in many developmental processes and in responses to various abiotic stresses, playing pivotal roles in plant adaptation. Deep sequencing technology was chosen to determine the small RNA transcriptome of Saccharum sp cultivars grown on saline conditions. We constructed four small RNAs libraries prepared from plants grown on hydroponic culture submitted to 170 mM NaCl and harvested after 1 h, 6 hs and 24 hs. Each library was sequenced individually and together generated more than 50 million short reads. Ninety-eight conserved miRNAs and 33 miRNAs* were identified by bioinformatics. Several of the microRNA showed considerable differences of expression in the four libraries. To confirm the results of the bioinformatics-based analysis, we studied the expression of the 10 most abundant miRNAs and 1 miRNA* in plants treated with 170 mM NaCl and in plants with a severe treatment of 340 mM NaCl. The results showed that 11 selected miRNAs had higher expression in samples treated with severe salt treatment compared to the mild one. We also investigated the regulation of the same miRNAs in shoots of four cultivars grown on soil treated with 170 mM NaCl. Cultivars could be grouped according to miRNAs expression in response to salt stress. Furthermore, the majority of the predicted target genes had an inverse regulation with their correspondent microRNAs. The targets encode a wide range of proteins, including transcription factors, metabolic enzymes and genes involved in hormone signaling, probably assisting the plants to develop tolerance to salinity. Our work provides insights into the regulatory functions of miRNAs, thereby expanding our knowledge on potential salt-stressed regulated genes.