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Meta-analyses and theory show that with rising atmospheric [CO 2 ], Rubisco has become the greatest limitation to light-saturated leaf CO 2 assimilation rates ( A sat ) in C 4 crops. So would transgenically increasing Rubisco increase A sat and result in increased productivity in the field? Here, we successfully overexpressed the Rubisco small subunit ( RbcS ) with Rubisco accumulation factor 1 ( Raf1 ) in both sorghum and sugarcane, resulting in significant increases in Rubisco content of 13 to 25% and up to 90% respectively. A sat increased 12 to 15% and Rubisco enzyme activity ~40% in three independent transgenic events of both species. Sorghum plants also showed increased speeds of photosynthetic induction and decreased bundle sheath leakiness. These improvements translated into average increases of 15.5% in biomass in field-grown sorghum and a 37 to 81% increase in greenhouse-grown sugarcane. This suggests a potential opportunity to achieve substantial increases in productivity of this key economically important clade of C 4 crops, future proofing their value under global atmospheric change., Competing Interests: Competing interests statement:The authors declare no competing interest.

Plant growth-promoting rhizobacteria (PGPR) are promising candidates that enhance plant growth under stressful conditions. In this study, 10 bacterial isolates were screened for their IAA production, among them JTB1 and MT22 isolates were selected which produced high IAA levels under 10% PEG and 2% NaCl stress. The isolates showed a high capacity for phosphate solubilization and ACC deaminase activity. Phytogenic analysis showed that the isolate belonged to Bacillus megaterium species JTB1 and MT22. Application of JTB1, MT22, and their consortia as PGPR significantly promoted root development and sugarcane growth under moderate and severe drought stress. Sugarcane growth promotion resulted from the retardation of reactive oxygen species (ROS) synthesis, malondialdehyde (MDA), electrolyte leakage, and cell damage by increasing antioxidant scavenging systems, such as catalase (CAT) and ascorbate peroxidase (APX), owing to PGPR inoculation under drought stress. Inoculation with PGPR resulted in increased auxin transporter expression, which modulated the increase in photosynthetic gene expression of RBC-L, PEPC, SPS in sugarcane under drought stress. The application of JTB1, MT22, or their consortia seemed to have similar effects on all observed parameters. Collectively, these results indicated that inoculation with PGPR enhanced root development and increased the antioxidant system and photosynthetic activity, which promoted sugarcane growth under drought stress., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Nitrogen (N) is an essential element for plant growth and development. Identifying functional gene loci associated with nitrogen absorption and utilization in sugar cane can facilitate the development of nutrient-efficient sugar cane varieties. In this study, sugar cane seedlings were subjected to normal and low nitrogen stress treatments within a hydroponic system for the identification of candidate genes related to six root-associated traits using a diversity population of 297 accessions. A total of 262 single nucleotide polymorphisms were found to be significantly associated with the mutation, including 30 stable or pleiotropic loci. The integration of genome-wide association studies and differential gene analysis from RNA sequencing (RNA-seq) identified five key candidate genes. Overexpression of one of them, ScMYB-CC gene, in Arabidopsis significantly affected root growth and development. In summary, the findings of this study provide valuable genetic resources for the breeding of nitrogen-efficient sugar cane varieties.

Sugarcane is a crop that accumulates sucrose with high photosynthesis efficiency. Therefore, the feedback regulation of sucrose on photosynthesis is crucial for improving sugarcane yield. Saccharum spontaneum and Saccharum officinarum are the two founding Saccharum species for modern sugarcane hybrids. S. spontaneum exhibits a higher net photosynthetic rate but lower sucrose content than S. officinarum. However, the mechanism underlying the negative feedback regulation of photosynthesis by sucrose remains poorly understood. This study investigates the effects of exogenous sucrose treatment on S. spontaneum and S. officinarum. Exogenous sucrose treatment increases sucrose content in the leaf base but inhibits photosynthetic efficiency and the expression of photosynthesis-related pathway genes (including RBCS and PEPC) in both species. However, gene expression patterns differed significantly, with few differentially expressed genes (DEGs) shared between the two species, indicating a differential response to exogenous sucrose. The expression networks of key genes involved in sugar metabolism, sugar transport, and PEPC and RBCS showed divergence between two species. Additionally, DEGs involved in the pentose phosphate pathway and the metabolism of alanine, aspartate, and glutamate metabolism were uniquely enriched in S. spontaneum, potentially contributing to the differential changes in sucrose content in the tip between the two species. We propose a model of the mechanisms underlying the negative feedback regulation of photosynthesis by sucrose in the leaves of S. spontaneum and S. officinarum. Our findings enhance the understanding of sucrose feedback regulation on photosynthesis and provide insights into the divergent molecular mechanisms of sugar accumulation in Saccharum., (© 2025 Society for Experimental Biology and John Wiley & Sons Ltd.)

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.)

Background: Glyphosate is an extensively employed herbicide in agriculture, specifically for sugarcane cultivation. The situation is different with the extensive physiological and genetic effects exerted by this herbicide on a range of plant species, including sugarcane, whose model basis is still poorly characterized, although its primary mode of action, which acts on the EPSPS enzyme in the shikimic acid pathway, is completely elucidated. The current study was aimed at investigating the stability of glyphosate formulation, molecular interactions of glyphosate formulation with rbcL enzyme associated with chlorophyll metabolism, and its effects on varieties of sugarcane., Methods and Results: The stability of a ground-up glyphosate formulation was assessed under accelerated storage conditions. Molecular docking was performed to analyze interactions between glyphosate derivatives and rbcL. Two sugarcane varieties (G84-47 and GT54-9) were treated with increasing glyphosate concentrations (0.2, 0.4, and 0.8 mg L -1 ) to evaluate effects on chlorophyll content, plant height, herbicide tolerance, and rbcL expression. The formulation showed good stability with minor degradation (47.55-47.14%). N-Nitrosoglyphosate and 2-(phosphonomethylamino)acetic acid exhibited favorable binding affinities with rbcL. Glyphosate treatment reduced chlorophyll content and plant height dose-dependently, with G84-47 showing higher sensitivity. GT54-9 demonstrated higher herbicide tolerance in survival analysis. rbcL expression remained stable in G84-47 but was significantly upregulated in GT54-9 under high herbicide stress., Conclusions: This study reveals genetic variability in sugarcane responses to glyphosate, with variety-specific mechanisms underlying stress adaptation. The inducible rbcL expression in tolerant varieties provides insights into herbicide resistance mechanisms. These findings can inform marker-assisted breeding and improve agronomic practices for enhanced sugarcane cultivation resilience and sustainability., Competing Interests: Declarations. Ethics approval and consent to participate: There is no ethical issue in this research. Competing interests: The authors declare no competing interests. Conflict of interest: There is no conflict of interest among the authors., (© 2025. The Author(s), under exclusive licence to Springer Nature B.V.)

The DHHC domain genes are crucial for protein lipid modification, a key post-translational modification influencing membrane targeting, subcellular trafficking, and protein function. Despite their significance, the DHHC gene family in Saccharinae remains understudied. Here, we identified 32 (110 alleles), 28, 53, and 48 DHHC genes in Saccharum spontaneum Np-X, Erianthus rufipilus, Miscanthus sinensis, and Miscanthus lutarioriparius, respectively. Collinearity analysis uncovered the loss of two M. lutarioriparius genes, homologues of EruDHHC1C and EruDHHC3A. Phylogenetic and classification analyses categorized DHHC family members into six subgroups (A-F). Ka/Ks ratio analysis indicated that gene duplication in these species was primarily driven by whole-genome duplication (WGD) and dispersed duplication (DSD), with DHHC genes evolving under strong purifying selection. Gene expression and trait correlation analysis revealed a significant negative correlation between SspDHHC28A expression in S. spontaneum and sucrose content, suggesting a role in photosynthesis product transport during rapid growth. This study deepens our understanding of the DHHC gene family's functional dynamics and evolutionary path in Saccharinae, laying a foundation for future research., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

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.

Main Conclusion: Significant changes in the proteome highlight essential metabolic adaptations for development and oxidative signaling induced by the treatment of young sugarcane plants with hydrogen peroxide. These adaptations suggest that hydrogen peroxide acts not only as a stressor but primarily as a signaling molecule, triggering specific metabolic pathways that regulate growth and plant resilience. Sugarcane is a crucial crop for sugar and ethanol production, often influenced by environmental signals. Hydrogen peroxide (H 2 O 2 ) is increasingly recognized as an important signaling molecule that regulates plant development and adaptation. In this study, two-month-old sugarcane plants were treated with varying concentrations of H 2 O 2 to investigate how this molecule acts as a signal at the cellular, biochemical, and proteomic levels. Antioxidant enzyme activity exhibited fluctuations, suggesting a dynamic response to oxidative signaling. Lipid peroxidation, observed through TBARs and scanning electron microscopy, highlighted early membrane modifications. Proteomic analysis (ProteomeXchange PXD048142) identified 2,699 proteins, with 155 showing significant expression changes in response to H 2 O 2 signaling. Bioinformatics, including Principal Component Analysis, revealed distinct proteomic profiles in roots and leaves, indicating tissue-specific metabolic reprogramming. Functional annotation through Gene Ontology and KEGG pathway enrichment showed that oxidative signaling led to the repression of photosynthesis-related pathways in leaves, while promoting pathways related to protein processing, glycolysis, and carbon metabolism in roots. Additionally, bioinformatic tools identified proteins involved in amino acid metabolism, the TCA cycle, and carbohydrate metabolism as critical components of sugarcane's adaptive signaling response. The data suggest that sugarcane plants responded to oxidative signals by adjusting their metabolic networks, promoting sustained development and potential pathways for targeted plant breeding., Competing Interests: Declarations. Ethics approval, guidelines and consent to participate: Not applicable. Consent for publication: The authors agree to the publication. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Background: Sugarcane is a crucial crop for both sugar and bioethanol production. The nobilization breeding and utilization of wild germplasm have significantly enhanced its productivity. However, the pollen sterility in Saccharum officinarum restricts its role to being a female parent in crosses with Saccharum spontaneum during nobilization breeding, resulting in a narrow genetic basis for modern sugarcane cultivars. Mitochondria, often referred to as the intracellular "energy factories", provide energy for plant life activities, and are also implicated in cytoplasmic male sterility (CMS)., Results: We performed mitochondrial genome assembly and structural analysis of two Saccharum founding species. We discovered that the proportions of repeat sequences are the primary factor contributing to the variations in mitochondrial genome structure and size between the two Saccharum species. Heterologous expression of the mitochondrial chimeric gene ORF113, which is highly expressed in male-sterile S. officinarum flowers, significantly inhibits growth and ATP synthesis in yeast cells, making it a key candidate CMS-related gene in sugarcane. Furthermore, we developed two co-dominant simple sequence repeat (SSR) markers based on the mitochondrial genome, which can effectively distinguish the cytoplasmic types of the two Saccharum species., Conclusion: In this study, we identified structural variants and developed SSR molecular markers in the mitochondrial genomes of both S. officinarum and S. spontaneum. We also identified a novel mitochondrial chimeric ORF as a key candidate CMS-related gene. These findings offer valuable insights into variety identification, genetic resource development, and cross-breeding strategies in sugarcane., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Collection of plant material: The collection of plant material complies with relevant institutional, national, and international guidelines and legislation. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Mosaic disease is one of the predominant viral diseases affecting sugarcane plants worldwide. In China, it is mainly caused by the sorghum mosaic virus (SrMV). Exogenous silicon (ESi) has emerged as a promising inducer of plant disease resistance. This study aims to elucidate the mechanistic effects of exogenous silicon on enhancing the resistance to SrMV in sugarcane. ESi was applied to the soil at different rates (15 g per barrel -1 , 30 g per barrel -1 , and 45 g per barrel -1 ), leading to a significant reduction in disease incidence (26.66-82.22%) compared to non-silicon-treated plants. Silicon application (15 g ESi barrel -1 , 30 g ESi barrel -1 ) mitigated SrMV inhibition of sugarcane growth, including plant height, stem diameter, and leaf area, while improving photosynthesis, including stomatal conductance, intercellular CO 2 concentration, net photosynthetic rate, and transpiration rate. Additionally, silicon mitigates SrMV-induced degradation of chlorophyll a and b and carotenoid content, alongside heightened activities of superoxide dismutase, peroxidase, and catalase, and decreased content of malondialdehyde and hydrogen peroxide in sugarcane leaves. The ultrastructural analysis revealed silicon's capacity to reduce SrMV accumulation within sugarcane mesophyll cells while preserving chloroplast integrity. Additionally, silicon application increases SA content in sugarcane leaves and upregulates the expression of key SA pathway genes (PAL, PR1, NPR1). These findings suggest that silicon may contribute to sugarcane resistance to SrMV by potentially influencing antioxidant enzyme activity, ROS production, and SA pathway genes., (© 2025 Scandinavian Plant Physiology Society.)

Modern sugarcane, a highly allo-autopolyploid organism, has a very complex genome. In the present study, the karyotype and genome architecture of modern sugarcane were investigated, resulting in a genome assembly of 97 chromosomes (8.84 Gb). The allopolyploid genome was divided into subgenomes from Saccharum officinarum (So h ) and S. spontaneum (Ss h ), with So h dominance in the Saccharum hybrid (S. hybrid). Genome shock affected transcriptome dynamics during allopolyploidization. Analysis of an inbreeding population with 192 individuals revealed the underlying genetic basis of transgressive segregation. Population genomics of 310 Saccharum accessions clarified the breeding history of modern sugarcane. Using the haplotype-resolved S. hybrid genome as a reference, genome-wide association studies identified a potential candidate gene for sugar content from S. spontaneum. These findings illuminate the complex genome evolution of allopolyploids, offering opportunities for genomic enhancements and innovative breeding strategies for sugarcane., Competing Interests: Competing interests: The authors declare no competing interests., (© 2025. The Author(s), under exclusive licence to Springer Nature America, Inc.)

The agricultural sugarcane residues, bagasse and straws, can be used for second-generation ethanol (2GE) production by the cellulose conversion into glucose (saccharification). However, the lignin content negatively impacts the saccharification process. This polymer is mainly composed of guaiacyl (G), hydroxyphenyl (H), and syringyl (S) units, the latter formed in the ferulate 5-hydroxylase (F5H) branch of the lignin biosynthesis pathway. We have generated transgenic lines overexpressing ShF5H1 under the control of the C4H (cinnamate 4-hydroxylase) rice promoter, which led to a significant increase of up to 160% in the S/G ratio and 63% in the saccharification efficiency in leaves. Nevertheless, the content of lignin was unchanged in this organ. In culms, neither the S/G ratio nor sucrose accumulation was altered, suggesting that ShF5H1 overexpression would not affect first-generation ethanol production. Interestingly, the bagasse showed a significantly higher fiber content. Our results indicate that the tissue-specific manipulation of the biosynthetic branch leading to S unit formation is industrially advantageous and has established a foundation for further studies aiming at refining lignin modifications. Thus, the ShF5H1 overexpression in sugarcane emerges as an efficient strategy to improve 2GE production from straw.

Sugarcane ( Saccharum spp.) is globally considered an important crop for sugar and biofuel production. During sugarcane production, the heavy reliance on chemical nitrogen fertilizer has resulted in low nitrogen use efficiency (NUE) and high loss. Up until now, there has been extensive research on the transcriptomic dynamics during sugarcane response to low nitrogen (LN) stress. However, the specific contribution of S. spontaneum to the NUE of modern sugarcane remains unclear. In the present study, the comparative transcriptome analysis of two contrasting sugarcane cultivars in response to nitrogen deficiency was performed via the combination of genomes of S. spontaneum and S. officinarum . Sub-genome analysis indicated that S. spontaneum supports the high NUE of modern sugarcane by providing genes related to nitrogen and carbohydrate metabolism, photosynthesis, and amino acid metabolism. Additionally, the key genes involved in nitrogen metabolism from the S. spontaneum were successfully identified through weighted gene co-expression network analyses (WGCNA), and a high-affinity nitrate transporter named ScNRT2.3 was subsequently cloned. Heterogeneous expression of the ScNRT2.3 , a cell membrane-localized protein, could enhance the growth of Arabidopsis under low nitrate conditions. Furthermore, a conserved protein module known as NAR2.1/NRT2.3 was shown to regulate the response to LN stress in sugarcane roots through molecular interaction. This work helps to clarify the contribution of S. spontaneum to the NUE in modern sugarcane, and the function of the ScNRT2.3 in sugarcane.

Background: Drought is one of main critical factors that limits sugarcane productivity and juice quality in tropical regions. The unprecedented changes in climate such as monsoon failure, increase in temperature and other factors warrant the need for development of stress tolerant cultivars to sustain sugar production. Plant Nuclear factor (NF-Y) is one of the major classes of transcription factors that have a major role in plant development and abiotic stress response. In our previous studies, we found that under drought conditions, the nuclear factor NF-YB2 was highly expressed in Erianthus arundinaceus, an abiotic stress tolerant wild genus of Saccharum species. In this study, the coding sequence of NF-YB2 gene was isolated from Erianthus arundinaceus and overexpressed in sugarcane to develop drought tolerant lines. RESULTS : EaNF-YB2 overexpressing sugarcane (OE) lines had higher relative water content, chlorophyll content and photosynthetic efficiency compared to non-transgenic (NT) control. In addition, overexpressing lines had higher activity of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR), and higher proline content, lower malondialdehyde (MDA) and peroxide (H 2 O 2 ) contents. The expression studies revealed that EaNF-YB2 expression was significantly higher in OE lines than NT control under drought stress. The OE lines had an elevated expression of abiotic stress responsive genes such as BRICK, HSP 70, DREB2, EDH45, and LEA3. The morphological analysis revealed that OE lines exhibited less wilting than NT under drought conditions., Conclusion: This study provides insights into the role of the EaNF-YB2 gene in drought tolerance in sugarcane. Based on the findings of this study, the EaNF-YB2 gene can be potentially exploited to produce drought tolerant sugarcane cultivars to sustain sugarcane production under water deficit conditions., Competing Interests: Declarations. Ethics approval and consent to participate: The Saccharum spp. Hybrid (Co 86032) used in this study was developed at the ICAR-Sugarcane Breeding Institute, Coimbatore, India, a government-funded institute. The experimental material involved follows institutional, national, and international guidelines and legislation. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Key Message: A total of 24 genes of vacuolar H + -translocating pyrophosphatases H + -PPases (VPP) genes were identified in Saccharum spontaneum AP85-441 and the ScVPP1-overexpressed Arabidopsis plants conferred salt tolerance. The vital role of vacuolar H + -translocating pyrophosphatases H + -PPases (VPP) genes involved in plants in response to abiotic stresses. However, the understanding of VPP functions in sugarcane remained unclear. In this study, a total of 24 VPP genes (SsaVPP1-SsaVPP24) were identified in the Saccharum spontaneum genome of haploid clone AP85-441. These genes were distributed in two phylogenetic groups. The SsaVPPs displayed diverse physio-chemical and gene structure attributes. The SsaVPP family genes have expanded through segmental duplication (20 gene pairs) rather than tandem duplication. A full-length cDNA of ScVPP1 was cloned from the sugarcane cultivar ROC22 and shared 99.48% sequence identity (amino acid) with homologous gene SsaVPP21 from AP85-441. In ROC22, the ScVPP1 gene was considerably upregulated by NaCl and ABA treatments among leaf, root, and stem tissues, while this gene was exclusively upregulated in the root with PEG treatment. Under NaCl and ABA stresses, yeast cells transfected by the ScVPP1 plasmid showed distinct growth rates compared to control yeast cells transfected by the empty vector. In transgenic Arabidopsis lines overexpressing ScVPP1, the seed gemination and survival rate were enhanced under NaCl treatment but not under ABA stress as compared to wild-type plants. These results suggested that the ScVPP1 gene conferred tolerance to slat and may be used as a salt resistance gene source for sugarcane breeding., Competing Interests: Declarations. Conflict of interest: The authors declare no conflicts of interest., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Elucidating the intricacies of the sugarcane genome is essential for breeding superior cultivars. This economically important crop originates from hybridizations of highly polyploid Saccharum species. However, the large size (10 Gb), high degree of polyploidy, and aneuploidy of the sugarcane genome pose significant challenges to complete genome sequencing, assembly, and annotation. One successful strategy for identifying candidate genes linked to agronomic traits, particularly those associated with sugar accumulation, leverages synteny and potential collinearity with related species., Results: In this study, we explored synteny between sorghum and sugarcane. Genes from a sorghum Brix QTL were used to screen bacterial artificial chromosome (BAC) libraries from two Brazilian sugarcane varieties (IACSP93-3046 and SP80-3280). The entire region was successfully recovered, confirming synteny and collinearity between the species. Manual annotation identified 51 genes in the hybrid varieties that were subsequently confirmed to be present in Saccharum spontaneum. This study employed a multifaceted approach to identify candidate genes for sugar accumulation, including retrieving the genomic region of interest, performing a gene-by-gene analysis, analyzing RNA-seq data for internodes from Saccharum officinarum and S. spontaneum accessions, constructing a coexpression network to examine the expression patterns of genes within the studied region and their neighbors, and finally identifying differentially expressed genes (DEGs)., Conclusions: This comprehensive approach led to the discovery of three candidate genes potentially involved in sugar accumulation: an ethylene-responsive transcription factor (ERF), an ABA 8'-hydroxylase, and a prolyl oligopeptidase (POP). These findings could be valuable for identifying additional candidate genes for other important agricultural traits and directly targeting candidate genes for further work in molecular breeding., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

GRAS gene family plays multifunctional roles in plant growth, development, and resistance to various biotic and abiotic stresses, belonging to the plant-specific transcription factor (TF) family. In this study, a genome-wide survey and systematic analysis of the GRAS family in cultivated hybrid sugarcane ZZ1 ( Saccharum officinarum ) with economic and industrial importance was carried out. We identified 747 GRAS genes with complete structural domains and classified these into 11 subfamilies by phylogenetic analyses, exhibiting a diverse range of molecular weight and isoelectric points, thereby indicating a broad structural and functional spectrum. Analysis of Protein motif and gene structure revealed a conserved yet variable arrangement of motifs within the GRAS TFs, suggesting its potential for diverse functional roles. Furthermore, the identification of numerous cis-regulatory elements by GRAS TFs promoter sequence analysis, implying their complex regulation in response to environmental and physiological signals. Tertiary structure predictions analyses using AlphaFold3 highlighted the structural flexibility and conservation within the GRAS family, with disordered regions potentially contributing to their functional versatility. Weighted Gene Co-expression Network Analysis (WGCNA) provided insights into the potential roles of ShGRAS21A in sugarcane's response to smut infection. This comprehensive investigation of the GRAS family in ZZ1 not only uncovers their structural diversity but also sheds light on their potential regulatory roles in plant growth, development, and stress response. The findings contribute to a deeper understanding of GRAS TFs functions and lay the groundwork for future studies on their role in sugarcane improvement and disease resistance.

The efficacy of generalized sugarcane yield prediction models holds significant implications for global food security. Given that machine learning algorithms often surpass the precision of remote sensing technology, further exploration of machine learning algorithms in the development of sugarcane yield prediction models is imperative. In this study, we employed six key phenotypic traits of sugarcane, specifically plant height, stem diameter, third-node length (internode length), leaf length, leaf width, and field brix, along with eight machine learning methods: logistic regression, linear regression, K-Nearest Neighbors (KNN), Support Vector Machine (SVM), Backpropagation Neural Network (BPNN), Decision Tree, Random Forest, and the XGBoost algorithm. The aim was to establish an intelligent model ensemble for predicting two crucial phenotypic characteristics-stem diameter and plant height-that determine sugarcane yield, ultimately enhancing the overall yield.The experimental findings indicate that the XGBoost algorithm outperforms the other seven algorithms in predicting these significant phenotypic traits of sugarcane. Furthermore, an analysis of the sugarcane intelligent prediction model's performance under a specialized data environment, incorporating self-prepared data, reveals that the XGBoost algorithm exhibits greater stability. Notably, the data pertaining to these crucial phenotypic traits have a profound impact on the efficacy of the intelligent models. The research demonstrates that a sugarcane yield prediction model ensemble, incorporating multiple intelligent algorithms, can accurately forecast stem diameter and plant height, thereby predicting sugarcane yield. Additionally, this approach, combined with the principles of sugarcane cross-breeding, provides a valuable reference for the artificial breeding of new sugarcane varieties that excel in stem diameter and plant height, bridging a research gap in indirect yield prediction through sugarcane phenotypic traits., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Sun et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The MADS-box transcription factor (TF) gene family is pivotal in various aspects of plant biology, particularly in growth, development, and environmental adaptation. It comprises Type I and Type II categories, with the MIKC-type subgroups playing a crucial role in regulating genes essential for both the vegetative and reproductive stages of plant life. Notably, MADS-box proteins can influence processes such as flowering, fruit ripening, and stress tolerance. Here, we provide a comprehensive overview of the structural features, evolutionary lineage, multifaceted functions, and the role of MADS-box TFs in responding to biotic and abiotic stresses. We particularly emphasize their implications for crop enhancement, especially in light of recent advances in understanding the impact on sugarcane ( Saccharum spp.), a vital tropical crop. By consolidating cutting-edge findings, we highlight potential avenues for expanding our knowledge base and enhancing the genetic traits of sugarcane through functional genomics and advanced breeding techniques. This review underscores the significance of MADS-box TFs in achieving improved yields and stress resilience in agricultural contexts, positioning them as promising targets for future research in crop science.

In agronomically important C4 grasses, efficient CO2 delivery to Rubisco is facilitated by NADP-malic enzyme (C4NADP-ME), which decarboxylates malate in bundle sheath cells. However, understanding the molecular regulation of the C4NADP-ME gene in sugarcane (Saccharum spp.) is hindered by its complex genetic background. Enzymatic activity assays demonstrated that decarboxylation in sugarcane Saccharum spontaneum predominantly relies on the NADP-ME pathway, similar to sorghum (Sorghum bicolor) and maize (Zea mays). Comparative genomics analysis revealed the recruitment of 8 core C4 shuttle genes, including C4NADP-ME (SsC4NADP-ME2), in the C4 pathway of sugarcane. Contrasting to sorghum and maize, the expression of SsC4NADP-ME2 in sugarcane is regulated by different transcription factors (TFs). We propose a gene regulatory network for SsC4NADP-ME2, involving candidate TFs identified through gene coexpression analysis and yeast 1-hybrid experiment. Among these, ABA INSENSITIVE5 (ABI5) was validated as the predominant regulator of SsC4NADP-ME2 expression, binding to a G-box within its promoter region. Interestingly, the core element ACGT within the regulatory G-box was conserved in sugarcane, sorghum, maize, and rice (Oryza sativa), suggesting an ancient regulatory code utilized in C4 photosynthesis. This study offers insights into SsC4NADP-ME2 regulation, crucial for optimizing sugarcane as a bioenergy crop., Competing Interests: Conflict of interest statement. The authors declare no competing interests., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Drought stress is a common hazard faced by sugarcane growth, and utilizing microorganisms to enhance plant tolerance to abiotic stress has become an important method for sustainable agricultural development. Several studies have demonstrated that Streptomyces chartreuses WZS021 improves sugarcane tolerance to drought stress. However, the molecular mechanisms underlying tolerance at the transcriptional and metabolomic levels remain unclear. We comprehensively evaluated the physiological and molecular mechanisms by which WZS021 enhances drought tolerance in sugarcane, by performing transcriptome sequencing and non-targeted metabolomics; and examining rhizosphere soil properties and plant tissue antioxidant capacity. WZS021 inoculation improved the rhizosphere nutritional environment (AP, ammonia, OM) of sugarcane and enhanced the antioxidant capacity of plant roots, stems, and leaves (POD, SOD, CAT). Comprehensive analyses of the transcriptome and metabolome revealed that WZS021 mainly affects plant drought tolerance through phenylalanine metabolism, plant hormone signal transduction, and flavonoid biosynthesis pathways. The drought tolerance signaling molecules mediated by WZS021 include petunidin, salicylic acid, α-Linoleic acid, auxin, geranylgeraniol and phenylalanine, as well as key genes related to plant hormone signaling transduction (YUCCA, amiE, AUX, CYPs, PAL, etc.). Interestingly, inoculation with WZS021 during regular watering induces a transcriptome-level response to biological stress in sugarcane plants. This study further elucidates a WZS021-dependent rhizosphere-mediated regulatory mechanism for improving sugarcane drought tolerance, providing a theoretical basis for increasing sugarcane production capacity., Competing Interests: Declaration of competing interest All authors declare that there is no conflict of interest., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Key Message: Candidate male sterility genes were identified in sugarcane, which interacts with kinase-related proteins, transcription factors, and plant hormone signaling pathways to regulate stamen and anther development. Saccharum officinarum is a cultivated sugarcane species that its predominant feature is high sucrose content in stems. Flowering is necessary for breeding new cultivars but will terminate plant growth and reduce sugar yield. The wild sugarcane species Saccharum spontaneum has robust and viable pollen, whereas most S. officinarum accessions are male sterile, which is a desirable trait of a maternal parent in sugarcane breeding. To study male sterility and related regulatory pathways in sugarcane, we carried out RNAseq using flowers in different developmental stages between male-sterile S. officinarum accession 'LA Purple' and fertile S. spontaneum accession 'SES208'. Gene expression profiles were used to detect how genes are differentially expressed between male sterile and fertile flowers and to identify candidate genes for male sterility. Weighted gene correlation networks analysis (WGCNA) was conducted to investigate the regulatory networks. Transcriptomic analyses showed that 988 genes and 2888 alleles were differentially expressed in S. officinarum compared to S. spontaneum. Ten differentially expressed genes and thirty alleles were identified as candidate genes and alleles for male sterility in sugarcane. The gene Sspon.03G0007630 and two alleles of the gene Sspon.08G0002270, Sspon.08G0002270-2B and Sspon.08G0014700-1A, were involved in the early stamen or carpel development stages, while the remaining genes were classified into the post-meiosis stage. Gibberellin, auxin, and jasmonic acid signaling pathways are involved in the stamen development in sugarcane. The results expanded our knowledge of male sterility-related genes in sugarcane and generated genomic resources to facilitate the selection of ideal maternal parents to improve breeding efficiency., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Key Message: A chitinase gene ScChiVII1 which is involved in defense against pathogen stress was characterized in sugarcane. Chitinases, a subclass of pathogenesis-related proteins, catalyze chitin hydrolysis and play a key role in plant defense against chitin-containing pathogens. However, there is little research on disease resistance analysis of chitinase genes in sugarcane, and the systematic identification of their gene families has not been reported. In this study, 85 SsChi and 23 ShChi genes, which were divided into 6 groups, were identified from the wild sugarcane species Saccharum spontaneum and Saccharum hybrid cultivar R570, respectively. Transcriptome analysis and real-time quantitative PCR revealed that SsChi genes responded to smut pathogen stress. The chitinase crude extracted from the leaves of transgenic Nicotiana benthamiana plants overexpressing ScChiVII1 (a homologous gene of SsChi22a) inhibited the hyphal growth of Fusarium solani var. coeruleum and Sporisorium scitamineum. Notably, the chitinase and catalase activities and the jasmonic acid content in the leaves of ScChiVII1 transgenic N. benthamiana increased after inoculation with F solani var. coeruleum, but the salicylic acid, hydrogen peroxide, and malondialdehyde contents decreased. Comprehensive RNA sequencing of leaves before (0 day) and after inoculation (2 days) revealed that ScChiVII1 transgenic tobacco enhanced plant disease resistance by activating transcription factors and disease resistance-related signaling pathways, and modulating the expression of genes involved in the hypersensitive response and ethylene synthesis pathways. Taken together, this study provides comprehensive information on the chitinase gene family and offers potential genetic resources for disease resistance breeding in sugarcane., Competing Interests: Declarations. Conflict of interest: The authors declare that there is no conflict of interest., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Sugarcane is an economically important polyploid crop whose genetic complexity and limited fertility poses a challenge for crop improvement programs. Gamma radiation-induced mutagenesis is an alternate approach for generating a diverse array of agronomically useful mutants, accelerating varietal development in a long-duration crop like sugarcane. To develop agronomically useful mutants of a commercial sugarcane genotype Co 99004, gamma ray induced in vitro mutagenesis was carried out. The phenotypic variants of Co 99004 in V1 generation could be categorized into five distinct phenotypically scorable classes, including three chlorophyll mutants (albina, chlorina and chlorina pigmented) and two green mutants like wild type control. SRAP marker analyses indicated distinct genomic variation among the phenotypic mutants and control plants, with the polymorphic information content (PIC) ranging from 0 to 0.472. Further, the phylogenetic dendrogram derived from the SRAP marker data grouped the mutants into four distinct clusters clearly differentiating the phenotypic classification. Sequencing of selected SRAP amplicons indicated deletion/insertion of gene specific fragments. Interestingly, the loss of chlorophyll in albina and chlorina mutants showed gamma irradiation-induced deletions in the gene encoding FAR1-RELATED SEQUENCE 5-like protein, which is involved in chlorophyll biosynthesis. GC-MS based metabolome profiling showed alteration in tetrapyrrole biosynthesis, MEP (Methylerythritol Phosphate), and fatty acid biosynthesis pathways, indicating a significant metabolic variation in the chlorophyll mutants. Further characterization of the genetically distinct, non-lethal green wild type mutants can lead to the identification of agronomically useful mutants. In addition, the loss-of-function chlorophyll mutants can serve as a good source for comparative genomics studies aimed at gene-trait association., 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 © 2024 Elsevier Ltd. All rights reserved.)

Genomic selection can enhance the rate of genetic gain of cane and sucrose yield in sugarcane (Saccharum L.), an important industrial crop worldwide. We assessed the predictive ability (PA) for six traits, such as theoretical recoverable sugar (TRS), number of stalks (NS), stalk weight (SW), cane yield (CY), sugar yield (SY), and fiber content (Fiber) using 20,451 single nucleotide polymorphisms (SNPs) with 22 statistical models based on the genomic estimated breeding values of 567 genotypes within and across five stages of the Louisiana sugarcane breeding program. TRS and SW with high heritability showed higher PA compared to other traits, while NS had the lowest. Machine learning (ML) methods, such as random forest and support vector machine (SVM), outperformed others in predicting traits with low heritability. ML methods predicted TRS and SY with the highest accuracy in cross-stage predictions, while Bayesian models predicted NS and CY with the highest accuracy. Extended genomic best linear unbiased prediction models accounting for dominance and epistasis effects showed a slight improvement in PA for a few traits. When both NS and TRS, which can be available as early as stage 2, were considered in a multi-trait selection model, the PA for SY in stage 5 could increase up to 0.66 compared to 0.30 with a single-trait model. Marker density assessment suggested 9091 SNPs were sufficient for optimal PA of all traits. The study demonstrated the potential of using historical data to devise genomic prediction strategies for clonal selection early in sugarcane breeding programs., (© 2024 The Author(s). The Plant Genome published by Wiley Periodicals LLC on behalf of Crop Science Society of America.)

Cane molasses, a sugar-rich agro-industrial byproduct, was used to enhance the production of ε-poly-l-lysine (ε-PL) in Streptomyces albulus PD-1 as a cost-effective carbon source. The sucrose metabolism pathway was engineered by heterologously expressing sucrose-6-phosphate hydrolase from Escherichia coli W. The optimization of various promoters identified the SP44 promoter, increasing the total sugar utilization rate by 2.76-fold compared with the ermEp* promoter. Additionally, adaptive laboratory evolution improved the total sugar utilization rate. The evolved strain achieved an ε-PL titer of 2.65 ± 0.15 g/L in flask experiments, increasing the ε-PL titer by 7.16-fold compared with the unevolved strain. Comparative transcriptomic analyses revealed that the enhanced tolerance of the evolved strain to high concentrations of cane molasses was primarily due to modifications in the sucrose metabolism pathways, microbial metabolism in heavy metals and phenols, and the amino acids transport and metabolic pathways. These changes enabled more efficient ε-PL production. During fed-batch fermentation in a 5-L fermentor using a concentration of 50 g/L cane molasses, the ε-PL titer reached 36.88 ± 0.62 g/L, and dry cell weight was 41.1 ± 1.0 g/L. This study illustrates that cane molasses is an economical carbon source for producing ε-PL on an industrial scale.

Sugarcane has been grown all around the world to meet sugar demands for industrial sector. The current sugar recovery percentage in sugarcane cultivars is dismally low which demands scientific efforts for improvements. Multiple approaches were adopted to enhance sugar contents in commercial sugarcane plants in contrast to conventional plant breeding methods. The exploitation of biotechnological methods and exploration of isomaltulose synthetic genes presented a promising solution to increase the existing low level of sugar recovery percentage in Saccharum officinarum L. Isomaltulose synthase gene II was employed and integrated into plant expression vector driven under the leaf and stem specific promoters terminated by nopaline synthase gene in a cloning strategy shown in the present study. Three gene constructs were developed in various combinations driven under promoters Zea mays ubiquitin and Cestrum Yellow Leaf Curl virus in the single and double combined stacked system. The transformation was executed in multiple formats with single transformed events, double promoter transformation events and triple construct stacked promoters in sugarcane induced calli via the particle gene gun. The transformation of ImSyGII in sugarcane genotype HSF-240 was confirmed by molecular gene analysis while expression quantification was determined through Real Time PCR. Furthermore, HPLC was also done to harvest the increased amounts of Isomaltulose in transgenic sugarcane juice. The present work upheld the enhanced ImSyGII expression in leaves owing to the exploitation of ubiquitin, while the Cestrum Yellow Leaf Curl virus promoter enhanced gene expression in sugarcane stems. The employment of three gene constructs collectively produced elite sugar lines producing more than 78% enhancements in whole sugar recovery percentage. The mature internode proved highly efficient and receptive regarding the production of isomaltulose. Quantifications and sugar contents evaluations upheld an increased Brix ratio of transgenic sugarcane lines than control lines., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Awan et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Stomata regulate CO2 and water vapor exchange between leaves and the atmosphere. Stomata are a target for engineering to improve crop intrinsic water use efficiency (iWUE). One example is by expressing genes that lower stomatal density (SD) and reduce stomatal conductance (gsw). However, the quantitative relationship between reduced SD, gsw, and the mechanisms underlying it is poorly understood. We addressed this knowledge gap using low-SD sugarcane (Saccharum spp. hybrid) as a case study alongside a meta-analysis of data from 10 species. Transgenic expression of EPIDERMAL PATTERNING FACTOR 2 from Sorghum bicolor (SbEPF2) in sugarcane reduced SD by 26-38% but did not affect gsw compared with the wild type. Further, no changes occurred in stomatal complex size or proxies for photosynthetic capacity. Measurements of gas exchange at low CO2 concentrations that promote complete stomatal opening to normalize aperture size between genotypes were combined with modeling of maximum gsw from anatomical data. These data suggest that increased stomatal aperture is the only possible explanation for maintaining gsw when SD is reduced. Meta-analysis across C3 dicots, C3 monocots, and C4 monocots revealed that engineered reductions in SD are strongly correlated with lower gsw (r2=0.60-0.98), but this response is damped relative to the change in anatomy., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

The objective of present review is to provide a scientific overview of sugarcane as a potential feedstock for biofuel and use of genome editing approach for improvement of industrial and agronomical traits in sugarcane. Sugarcane, a perennial tropical grass with a high biomass index, is a promising feedstock for bioethanol production, and its bagasse, rich in lignocellulosic material, serves as an ideal feedstock for producing second-generation bioethanol. To improve the conversion of sugarcane biomass into biofuels, developing varieties with improved biomass degradability and high biomass and sucrose content is essential. The complex genome architecture and earlier lack of sequence data hindered biotechnological advancements in sugarcane, but recent genome sequence updates offer new opportunities for sugarcane improvement. The first genetically modified sugarcane was developed in 1992 by Bower and Birch using microprojectile bombardment of embryogenic callus. Since then, transgenic techniques have rapidly evolved, leading to the advancement of genome editing technologies. Application of genome editing tools particularly CRISPR/Cas system has been successfully used in sugarcane for editing. Recently, multiple alleles of the magnesium chelatase and acetolactate synthase genes in sugarcane have been successfully edited through multiplexing. Additionally, CRISPR-edited sugarcane varieties with modified cell wall components and increased sucrose content for enhanced bioethanol production have been developed. At the end, the future of CRISPR-edited crops will depend on how well regulatory frameworks adapt to the rapidly evolving technology., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Aldehyde Dehydrogenases (ALDH), a group of enzymes, are associated with the detoxification of aldehydes, produced in plants during abiotic stress conditions. Salinity remains a pivotal abiotic challenge that poses a significant threat to cultivation and yield of sugarcane. In this study, an Aldehyde dehydrogenase gene (EaALDH7) from Erianthus arundinaceus was overexpressed in the commercial sugarcane hybrid cultivar Co 86032. The transgenic lines were evaluated at different NaCl concentrations ranging from 0 mM to 200 mM for various morpho-physiological and biochemical parameters. The control plants, subjected to salinity stress condition, exhibited morphological changes in protoxylem, metaxylem, pericycle and pith whereas the transgenic events were on par with plants under regular irrigation. The overexpressing (OE) lines showed less cell membrane injury and improved photosynthetic rate, transpiration rate, and stomatal conductance than the untransformed control plants under stress conditions. Elevated proline content, higher activity of enzymatic antioxidants such as sodium dismutase (SOD), catalase (CAT), glutathione reductase (GR) and ascorbate peroxidase (APX) and low level of malondialdehyde MDA and hydrogen peroxide (H 2 O 2 ) in the transgenic lines. The analysis of EaALDH7 expression revealed a significant upregulation in the transgenic lines compared to that of the untransformed control during salt stress conditions. The current study highlights the potentials of EaALDH7 gene in producing salinity-tolerant sugarcane cultivars., Competing Interests: Declaration of Competing Interest The authors have declared that no competing interests exist., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Sugarcane (Saccharum spp.) growth is regulated by intricate gene networks and hormone secretions, positively correlating with sugarcane yield. There is a rising interest in exploring how the candidate genes found in sugarcane respond to plant growth. In this study, we simulated a typical growth environment to obtain accurate phenotypic data and screened for potential genes associated with plant growth through transcriptomics. Compared to Saccharum GuiTang 42, the other variety Saccharum GuiTang 44 exhibited earlier germination, a higher emergence rate, thicker pseudostems, taller plants, and a more extensive root system. The middle buds formed the greatest number of roots, followed by the lower and upper buds. Indole-3-acetic acid (IAA) and jasmonic acid effectively promoted bud development, while abscisic acid and trans-zeatin exhibited negative correlations with sugarcane bud growth. Transcriptome data from the upper, middle, and lower buds revealed 24,158 differentially expressed genes in all three comparisons, with MAPK signaling emerging as a critical pathway. The photosynthesis-antenna protein pathway is vital for middle and lower bud development during root germination. Lastly, key gene modules related to differences in hormone content between the two varieties were defined through weighted correlation network analysis and identified. The module significantly associated with IAA was enriched in pathways such as Proteasome and Protein processing in the endoplasmic reticulum, and the upregulation of key genes involved in this gene module had a highly significant positive correlation with bud outgrowth combined with IAA secretion. In conclusion, we have elucidated the pathways of hormones during sugarcane growth and the interactions between IAA and critical genes. These in-depth findings may guide modern sugarcane breeding., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The SnRK1, hexokinase, and TORC1 (TOR, LST8, RAPTOR) are three pivotal kinases at the core of sugar level sensing, significantly impacting plant metabolism and development. We retrieved and analyzed protein sequences of these three kinase pathways from seven sugarcane transcriptome and genome datasets, identifying protein domains, phylogenetic relationships, sequence ancestry, and in silico expression levels. Additionally, we predicted HXK subcellular localization and assessed its enzymatic activity in sugarcane leaves and culms along development in the field. We retrieved 11 TOR, 23 RAPTOR, 55 LST8, 95 SnRK1α, 98 HXK, and 14 HXK-like putative full-length sequences containing all the conserved domains. Most of these transcripts seem to share a common origin with the three ancestral species of sugarcane: Saccharum officinarum, Saccharum spontaneum, and Saccharum barberi. We accessed the expression profile of sequences from one sugarcane transcriptome. We found the highest enzymatic activity of HXK in culms in the first month, which, at this stage, provides carbon (sucrose) and nitrogen (amino acids) for initial plant development. Our approach places novel sugar sensing sequences that work as a guideline for further research into the underlying signaling mechanisms and biotechnology applications in sugarcane., (© 2024. The Author(s).)

Sugarcane smut caused by Sporisorium scitamineum represents the most destructive disease in the sugarcane industry, causing host hormone disruption and producing a black whip-like sorus in the apex of the stalk. In this study, the gibberellin metabolic pathway was found to respond to S. scitamineum infection, and the contents of bioactive gibberellins were significantly reduced in the leaves of diseased plants. The gibberellin receptor gene ScGID1 was identified and significantly downregulated. ScGID1 localized in both the nucleus and cytoplasm and had the highest expression level in the leaves. Eight proteins that interact with ScGID1 were screened out using a yeast two-hybrid assay. Novel DELLA proteins named ScGAI1a and ScGA20ox2, key enzymes in GA biosynthesis, were both found to interact with ScGID1 in a gibberellin-independent manner. Transcription factor trapping with a yeast one-hybrid system identified 50 proteins that interacted with the promoter of ScGID1 , among which ScS1FA and ScPLATZ inhibited ScGID1 transcription, while ScGDSL promoted transcription. Overexpression of ScGID1 in transgenic Nicotiana benthamiana plants could increase plant height and promote flowering. These results not only contribute to improving our understanding of the metabolic regulatory network of sugarcane gibberellin but also expand our knowledge of the interaction between sugarcane and pathogens.

Calcineurin B-like proteins (CBLs) perceive calcium signals triggered by abiotic stress and interact with CBL-interacting protein kinases (CIPKs) to form a complex signal network. This study identified 21 SsCBL and 89 SsCIPK genes in Saccharum spontaneum, and 90 ScCBL and 367 ScCIPK genes in the sugarcane cultivar ZZ1. Phylogenetic analysis classified CBL genes into three groups and CIPK genes into twenty-five groups, with whole-genome duplication events promoting their expansion in sugarcane. RNA-seq analysis revealed their involvement in abiotic stress responses through ABA, JA, and SA pathways. Four ScCBLs and eight ScCIPKs were cloned from ZZ1. Three CBL-CIPK interactions were detected using a yeast two-hybrid system and Firefly luciferase complementation imaging, showing CBLs as membrane proteins and CIPKs as nuclear proteins. Spatial expression profiles indicate these genes are expressed in various tissues, with the highest expression in roots. Gene expression analyses suggested that CBL-CIPK signaling networks are involved in responses to drought, salt, and reactive oxygen species, possibly through Ca 2+ -induced hormone pathways. These findings establish three CBL-CIPK signaling networks responding to abiotic stress, providing a molecular basis for improving sugarcane stress resistance., 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 © 2024 Elsevier B.V. All rights reserved.)

Sugarcane (Saccharum spp. hybrid) is a prime feedstock for commercial production of biofuel and table sugar. Optimizing canopy architecture for improved light capture has great potential for elevating biomass yield. LIGULELESS1 (LG1) is involved in leaf ligule and auricle development in grasses. Here, we report CRISPR/Cas9-mediated co-mutagenesis of up to 40 copies/alleles of the putative LG1 in highly polyploid sugarcane (2n = 100-120, x = 10-12). Next generation sequencing revealed co-editing frequencies of 7.4%-100% of the LG1 reads in 16 of the 78 transgenic lines. LG1 mutations resulted in a tuneable leaf angle phenotype that became more upright as co-editing frequency increased. Three lines with loss of function frequencies of ~12%, ~53% and ~95% of lg1 were selected following a randomized greenhouse trial and grown in replicated, multi-row field plots. The co-edited LG1 mutations were stably maintained in vegetative progenies and the extent of co-editing remained constant in field tested lines L26 and L35. Next generation sequencing confirmed the absence of potential off targets. The leaf inclination angle corresponded to light transmission into the canopy and tiller number. Line L35 displaying loss of function in ~12% of the lg1 NGS reads exhibited an 18% increase in dry biomass yield supported by a 56% decrease in leaf inclination angle, a 31% increase in tiller number, and a 25% increase in internode number. The scalable co-editing of LG1 in highly polyploid sugarcane allows fine-tuning of leaf inclination angle, enabling the selection of the ideotype for biomass yield., (© 2024 The Author(s). Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Competing Interests: Conflict of interest statement. The authors declare no conflict of interest.

12-Oxo-phytodienoic acid reductases (OPRs) perform vital functions in plants. However, few studies have been reported in sugarcane (Saccharum spp.), and it is of great significance to systematically investigates it in sugarcane. Here, 61 ShOPRs, 32 SsOPRs, and 36 SoOPRs were identified from R570 (Saccharum spp. hybrid cultivar R570), AP85-441 (Saccharum spontaneum), and LA-purple (Saccharum officinarum), respectively. These OPRs were phylogenetically classified into four groups, with close genes similar structures. During evolution, OPR gene family was mainly expanded via whole-genome duplications/segmental events and predominantly underwent purifying selection, while sugarcane OPR genes may function differently in response to various stresses. Further, ScOPR2, a tissue-specific OPR, which was localized in cytoplasm and cell membrane and actively response to salicylic acid (SA), methyl jasmonate, and smut pathogen (Sporisorium scitamineum) stresses, was cloned from sugarcane. In addition, both its transient overexpression and stable overexpression enhanced the resistance of transgenic plants to pathogen infection, most probably through activating pathogen-associated molecular pattern/pattern-recognition receptor-triggered immunity, producing reactive oxygen species, and initiating mitogen-activated protein kinase cascade. Subsequently, the transmission of SA and hypersensitive reaction were triggered, which stimulated the transcription of defense-related genes. These findings provide insights into the function of ScOPR2 gene for disease resistance., (© 2024 Society for Experimental Biology and John Wiley & Sons Ltd.)

Sugarcane (Saccharum spp.)is an economically useful crop grown globally for sugar, ethanol and biofuel production. The crop is vulnerable to fungus Colletotrichum falcatum known to cause red rot disease. The pathogen hydrolyses stalk parenchyma cells where sucrose is accumulated resulting in upto 75% losses in sugar recovery. In this study, transgenic sugarcane having resistance against red rot was developed by introducing Trichoderma spp. endochitinase following Agrobacterium mediated transformation. The transgene introduction and expression in genetically modified plants were verified through qRT-PCR revealing upto 6-fold enhancement in endochitinase expression than non-transgenic plants. Hyperspectral Imaging of transgenic plants displayed altered leaf reflectance spectra and vegetative indices that were positively correlated with ransgene expression. The bioassay with virulent pathotypes of C. falcatumCF08 and CF13 known for epiphytotic occurrence resulted in identification of resistant plant Chit 3-13.The plants with higher reflectance also displayed improved disease resistance, implying their early classification into resistant/susceptible. The losses in sucrose content were minimized (up to 4-fold) in inoculated resistant plant Chit 3-13 as compared to susceptible non-transgenic plant, and a fewer pathogen hyphae were detected in vascular cells of the former through optical microscopy. The electron micrographs confirmed sucrose-filled stalk parenchyma cells in Chit 3-13; in contrast, cells of non-transgenic inoculated plant were depleted of sucrose. The active sites involved in cleaving 1-4 β-glycoside bonds of N-acetyl-d-glucosaminein the pathogen hyphal walls were detected through endochitinase protein structural modelling. The transgenic sugarcane is an important source for in trogressingred rot resistance in plant breeding programs., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Sharma et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Main Conclusion: The high intrinsic water-use efficiency of Erianthus may be due to the low abaxial stomatal density and the accumulation of leaf metabolites such as betaine and gamma-aminobutyric acid. Sugarcane is an important crop that is widely cultivated in tropical and subtropical regions of the world. Because drought is among the main impediments limiting sugarcane production in these regions, breeding of drought-tolerant sugarcane varieties is important for sustainable production. Erianthus arundinaceus, a species closely related to sugarcane, exhibits high intrinsic water-use efficiency (iWUE), the underlying mechanisms for which remain unknown. To improve the genetic base for conferring drought tolerance in sugarcane, in the present study, we performed a comprehensive comparative analysis of leaf gas exchange and metabolites in different organs of sugarcane and Erianthus under wet and dry soil-moisture conditions. Erianthus exhibited lower stomatal conductance under both conditions, which resulted in a higher iWUE than in sugarcane. Organ-specific metabolites showed gradations between continuous parts and organs, suggesting linkages between them. Cluster analysis of organ-specific metabolites revealed the effects of the species and treatments in the leaves. Principal component analysis of leaf metabolites confirmed a rough ordering of the factors affecting their accumulations. Compared to sugarcane leaf, Erianthus leaf accumulated more raffinose, betaine, glutamine, gamma-aminobutyric acid, and S-adenosylmethionine, which function as osmolytes and stress-response compounds, under both the conditions. Our extensive analyses reveal that the high iWUE of Erianthus may be due to the specific accumulation of such metabolites in the leaves, in addition to the low stomatal density on the abaxial side of leaves. The identification of drought-tolerance traits of Erianthus will benefit the generation of sugarcane varieties capable of withstanding drought stress., (© 2024. The Author(s).)

From the fluff generated during 2005, after the preliminary experiments (2005-2007), a promising clone G2005047 has been identified. It showed moderate resistance to red rot (3.6 on a 9-scale scoring system), less susceptibility to shoot borer (13.25%) and internode borers (25.35%), and resistance to woolly aphid (0%). In the Advanced Yield Trials (2008-2011), it showed advantages over check for cane yield (CY) (11.79%), commercial cane sugar percent (CCSP) (0.35%), and sugar yield (SY) (20.33%). To ascertain its large-scale cultivation suitability, it has experimented under adaptive research trials (2012-2014) at farmers' fields. It exhibited 18.04%, 1.27%, and 19.55% supremacy over the check Co 86032 for CY, CCSP, and SY respectively. The stability of G2005047 under salinity was ascertained through a multi-environment-based experiment (2015-2017). AMMI (Additive Main-effects and Multiplicative Interactions) and GGE (Genotype × Genotype-Environment interaction) biplots were utilized. ANOVA revealed that the genotypic variation exerted the most significant effect followed by genotype × environment interaction and environment. G2005047 had the highest mean values for yield and quality traits with minimal ASV (AMMI stability value) (2.38:CY; 0.57: CCSP; & 0.58:SY) indicating its good-yielding ability and stability. AMMI I, AMMI II, and GGE biplots confirmed the stability of G2005047. In the jaggery quality assessment trials (2018 and 2019), it yielded 37.1% increased jaggery over the check. Also, the clone G2005047, exhibited moderate resistance to red rot disease, less susceptibility to shoot borer (13.25%) and internode borer (25.35%), and resistance against sugarcane woolly aphid (SWA). Due to supremacy for yield, quality, better performance under salinized situations, and tolerance to disease and pests, the clone G2005047 was released as a variety CoG 7 in 2022., (© 2024. The Author(s).)

Sugarcane (Saccharum spp.) plays a crucial role in global sugar production; however, the efficiency of breeding programs has been hindered by its heterozygous polyploid genomes. Considering non-additive genetic effects is essential in genome prediction (GP) models of crops with highly heterozygous polyploid genomes. This study incorporates non-additive genetic effects and pedigree information using machine learning methods to track sugarcane breeding lines and enhance the prediction by assessing the degree of association between genotypes. This study measured the stalk biomass and sugar content of 297 clones from 87 families within a breeding population used in the Japanese sugarcane breeding program. Subsequently, we conducted analyses based on the marker genotypes of 33,149 single-nucleotide polymorphisms. To validate the accuracy of GP in the population, we first predicted the prediction accuracy of the best linear unbiased prediction (BLUP) based on a genomic relationship matrix. Prediction accuracy was assessed using two different cross-validation methods: repeated 10-fold cross-validation and leave-one-family-out cross-validation. The accuracy of GP of the first and second methods ranged from 0.36 to 0.74 and 0.15 to 0.63, respectively. Next, we compared the prediction accuracy of BLUP and two machine learning methods: random forests and simulation annealing ensemble (SAE), a newly developed machine learning method that explicitly models the interaction between variables. Both pedigree and genomic information were utilized as input in these methods. Through repeated 10-fold cross-validation, we found that the accuracy of the machine learning methods consistently surpassed that of BLUP in most cases. In leave-one-family-out cross-validation, SAE demonstrated the highest accuracy among the methods. These results underscore the effectiveness of GP in Japanese sugarcane breeding and highlight the significant potential of machine learning methods., (© 2024 The Author(s). The Plant Genome published by Wiley Periodicals LLC on behalf of Crop Science Society of America.)

Sugarcane is the main source of sugar worldwide, and 80% of the sucrose production comes from sugarcane. However, the genetic differentiation and basis of agronomic traits remain obscure. Here, we sequenced the whole-genome of 219 elite worldwide sugarcane cultivar accessions. A total of approximately 6 million high-quality genome-wide single nucleotide polymorphisms (SNPs) were detected. A genome-wide association study identified a total of 2198 SNPs that were significantly associated with sucrose content, stalk number, plant height, stalk diameter, cane yield, and sugar yield. We observed homozygous tendency of favor alleles of these loci, and over 80% of cultivar accessions carried the favor alleles of the SNPs or haplotypes associated with sucrose content. Gene introgression analysis showed that the number of chromosome segments from Saccharum spontaneum decreased with the breeding time of cultivars, while those from S. officinarum increased in recent cultivars. A series of selection signatures were identified in sugarcane improvement procession, of which 104 were simultaneously associated with agronomic traits and 45 of them were mainly associated with sucrose content. We further proposed that as per sugarcane transgenic experiments, ShN/AINV3.1 plays a positive role in increasing stalk number, plant height, and stalk diameter. These findings provide comprehensive resources for understanding the genetic basis of agronomic traits and will be beneficial to germplasm innovation, screening molecular markers, and future sugarcane cultivar improvement., (© 2024 Society for Experimental Biology and John Wiley & Sons Ltd.)

The NAC (NAM, ATAF, and CUC) is one of the largest transcription factor gene families in plants. In this study, 180, 141, and 131 NAC family members were identified from Saccharum complex, including S. officinarum, S. spontaneum, and Erianthus rufipilus. The Ka/Ks ratio of ATAF subfamily was all less than 1. Besides, 52 ATAF members from 12 representative plants were divided into three clades and there was only a significant expansion in maize. Surprisingly, ABA and JA cis-elements were abundant in hormonal response factor, followed by transcriptional regulator and abiotic stressor. The ATAF subfamily was differentially expressed in various tissues, under low temperature and smut pathogen treatments. Further, the ScATAF1 gene, with high expression in leaves, stem epidermis, and buds, was isolated. The encoded protein, lack of self-activation activity, was situated in the cell nucleus. Moreover, SA and JA stresses down-regulated the expression of this gene, while ABA, NaCl, and 4°C treatments led to its up-regulation. Interestingly, its expression in the smut susceptible sugarcane cultivars was much higher than the smut resistant ones. Notably, the colors presented slight brown in tobacco transiently overexpressing ScATAF1 at 1 d after DAB staining, while the symptoms were more obvious at 3 d after inoculation with Ralstonia solanacearum, with ROS, JA, and SA signaling pathway genes significantly up-regulated. We thus speculated ScATAF1 gene could negatively mediate hypersensitive reactions and produce ROS by JA and SA signaling pathways. These findings lay the groundwork for in-depth investigation on the biological roles of ATAF subfamily in sugarcane., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Background: The Nuclear Factor Y (NF-Y) transcription factor (TF) gene family plays a crucial role in plant development and response to stress. Limited information is available on this gene family in sugarcane., Objectives: To identify sugarcane NF-Y genes through bioinformatic analysis and phylogenetic association and investigate the expression of these genes in response to abiotic and biotic stress., Methods: Sugarcane NF-Y genes were identified using comparative genomics from functionally annotated Poaceae and Arabidopsis species. Quantitative PCR and transcriptome analysis assigned preliminary functional roles to these genes in response to water deficit, cold and African sugarcane borer (Eldana saccharina) infestation., Results: We identify 21 NF-Y genes in sugarcane. Phylogenetic analysis revealed three main branches representing the subunits with potential discrepancies present in the assignment of numerical names of some NF-Y putative orthologs across the different species. Gene expression analysis indicated that three genes, ShNF-YA1, A3 and B3 were upregulated and two genes, NF-YA4 and A7 were downregulated, while three genes were upregulated, ShNF-YB2, B3 and C4, in the plants exposed to water deficit and cold stress, respectively. Functional involvement of NF-Y genes in the biotic stress response were also detected where three genes, ShNF-YA6, A3 and A7 were downregulated in the early resistant (cv. N33) response to Eldana infestation whilst only ShNF-YA6 was downregulated in the susceptible (cv. N11) early response., Conclusions: Our research findings establish a foundation for investigating the function of ShNF-Ys and offer candidate genes for stress-resistant breeding and improvement in sugarcane., (© 2024. The Author(s).)

Brazil is the largest global producer of sugarcane and plays a significant role-supplier of sugar and bioethanol. However, diseases such as brown and orange rust cause substantial yield reductions and economic losses, due decrease photosynthesis and biomass in susceptible cultivars. Molecular markers associated with resistance genes, such as Bru1 (brown rust) and G1 (orange rust), could aid in predicting resistant genotypes. In this study, we sought to associate the phenotypic response of 300 sugarcane accessions with the genotypic response of Bru1 and G1 markers. The field trials were conducted in a randomized block design, and five six-month-old plants per plot were evaluated under natural disease conditions. Genotypic information about the presence or absence of Bru1 (haplotype 1) and G1 gene was obtained after extraction of genomic DNA and conventional PCR. Of the total accessions evaluated, 60.3% (181) showed resistance to brown rust in the field, and of these, 70.7% (128) had the Bru1 gene present. Considering the field-resistant accessions obtained from Brazilian breeding programs (116), the Bru1 was present in 77,6% of these accessions. While alternative resistance sources may exist, Bru1 likely confers enduring genetic resistance in current Brazilian cultivars. Regarding the phenotypic reaction to orange rust, the majority of accessions, 96.3% (288), were field resistant, and of these, 52.7% (152) carried the G1 marker. Although less efficient for predicting resistance when compared to Bru1, the G1 marker could be part of a quantitative approach when new orange rust resistance genes are described. Therefore, these findings showed the importance of Bru1 molecular markers for the early selection of resistant genotypes to brown rust by genetic breeding programs., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Oliveira et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Leaf scald, caused by Xanthomonas albilineans, is a severe disease affecting sugarcane worldwide. One of the most practical ways to control it is by developing resistant sugarcane cultivars. It is essential to identify genes associated with the response to leaf scald. A panel of 170 sugarcane genotypes was evaluated for resistance to leaf scald in field conditions for 2 years, followed by a 1-year greenhouse experiment. The phenotypic evaluation data showed a wide continuous distribution, with heritability values ranging from 0.58 to 0.84. Thirteen single nucleotide polymorphisms (SNPs) were identified, significantly associated with leaf scald resistance. Among these, eight were stable across multiple environments and association models. The candidate genes identified and validated based on RNA-seq and qRT-PCR included two genes that encode NB-ARC leucine-rich repeat (LRR)-containing domain disease-resistance protein. These findings provide a basis for developing marker-assisted selection strategies in sugarcane breeding programs., (© 2024. The Author(s).)

Key Message: We reported the graph-based mitochondrial genomes of three foundation species (Saccharum spontaneum, S. robustum and S. officinarum) for the first time. The results revealed pan-structural variation and evolutionary processes in the mitochondrial genomes within Saccharum. Saccharum belongs to the Andropogoneae, and cultivars species in Saccharum contribute nearly 80% of sugar production in the world. To explore the genomic studies in Saccharum, we assembled 15 complete mitochondrial genomes (mitogenome) of three foundation species (Saccharum spontaneum, S. robustum and S. officinarum) using Illumina and Oxford Nanopore Technologies sequencing data. The mitogenomes of the three species were divided into a total of eight types based on contig numbers and linkages. All mitogenomes in the three species encoded 51 unique genes, including 32 protein-coding, 3 ribosomal RNA (rRNA) and 16 transfer RNA (tRNA) genes. The existence of long and short-repeat-mediated recombinations in the mitogenome of S. officinarum and S. robustum was revealed and confirmed through PCR validation. Furthermore, employing comparative genomics and phylogenetic analyses of the organelle genomes, we unveiled the evolutionary relationships and history of the major interspecific lineages in Saccharum genus. Phylogenetic analyses of homologous fragments between S. officinarum and S. robustum showed that S. officinarum and S. robustum are phylogenetically distinct and that they were likely parallel rather than domesticated. The variations between ancient (S. sinense and S. barberi) and modern cultivated species (S. hybrid) possibly resulted from hybridization involving different S. officinarum accessions. Lastly, this project reported the first graph-based mitogenomes of three Saccharum species, and a systematic comparison of the structural organization, evolutionary processes, and pan-structural variation of the Saccharum mitogenomes revealed the differential features of the Saccharum mitogenomes., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The auxin/indoleacetic acid ( Aux/IAA ) family plays a central role in regulating gene expression during auxin signal transduction. Nonetheless, there is limited knowledge regarding this gene family in sugarcane. In this study, 92 members of the IAA family were identified in Saccharum spontaneum , distributed on 32 chromosomes, and classified into three clusters based on phylogeny and motif compositions. Segmental duplication and recombination events contributed largely to the expansion of this superfamily. Additionally, cis-acting elements in the promoters of SsIAAs involved in plant hormone regulation and stress responsiveness were predicted. Transcriptomics data revealed that most SsIAA expressions were significantly higher in stems and basal parts of leaves, and at nighttime, suggesting that these genes might be involved in sugar transport. QRT-PCR assays confirmed that cold and salt stress significantly induced four and five SsIAAs , respectively. GFP-subcellular localization showed that SsIAA23 and SsIAA12a were localized in the nucleus, consistent with the results of bioinformatics analysis. In conclusion, to a certain extent, the functional redundancy of family members caused by the expansion of the sugarcane IAA gene family is related to stress resistance and regeneration of sugarcane as a perennial crop. This study reveals the gene evolution and function of the SsIAA gene family in sugarcane, laying the foundation for further research on its mode of action.