UAV-based time-series phenotyping reveals the genetic basis of plant height in upland cotton.

Plant height (PH) is an important agronomic trait affecting crop architecture, biomass, resistance to lodging and mechanical harvesting. Elucidating the genetic governance of plant height is crucial because of the global demand for high crop yields. However, during the rapid growth period of plants the PH changes a lot on a daily basis, which makes it difficult to accurately phenotype the trait by hand on a large scale. In this study, an unmanned aerial vehicle (UAV)-based remote-sensing phenotyping platform was applied to obtain time-series PHs of 320 upland cotton accessions in three different field trials. The results showed that the PHs obtained from UAV images were significantly correlated with ground-based manual measurements, for three trials (R2 = 0.96, 0.95 and 0.96). Two genetic loci on chromosomes A01 and A11 associated with PH were identified by genome-wide association studies (GWAS). GhUBP15 and GhCUL1 were identified to influence PH in further analysis. We obtained a time series of PH values for three field conditions based on remote sensing with UAV. The key genes identified in this study are of great value for the breeding of ideal plant architecture in cotton.

Natural variation in Beauty Mark is associated with UV-based geographical adaptation in Gossypium species.

BACKGROUND: Anthocyanins, a class of specialized metabolites that are ubiquitous among plant species, have attracted a great deal of attention from plant biologists due to their chemical diversity. They confer purple, pink, and blue colors that attract pollinators, protect plants from ultraviolet (UV) radiation, and scavenge reactive oxygen species (ROS) to facilitate plant survival during abiotic stress. In a previous study, we identified Beauty Mark (BM) in Gossypium barbadense as an activator of the anthocyanin biosynthesis pathway; this gene also directly led to the formation of a pollinator-attracting purple spot. RESULTS: Here, we found that a single nucleotide polymorphism (SNP) (C/T) within the BM coding sequence was responsible for variations in this trait. Transient expression assays of BM from G. barbadense and G. hirsutum in Nicotiana benthamiana using luciferase reporter gene also suggested that SNPs in the coding sequence could be responsible for the absent beauty mark phenotype observed in G. hirsutum. We next demonstrated that the beauty mark and UV floral patterns are associated phenotypes and that UV exposure resulted in increased ROS generation in floral tissues; BM thus contributed to ROS scavenging in G. barbadense and wild cotton plants with flowers containing the beauty mark. Furthermore, a nucleotide diversity analysis and Tajima's D Test suggested that there have been strong selective sweeps in the GhBM locus during G. hirsutum domestication. CONCLUSIONS: Taken together, these results suggest that cotton species differ in their approaches to absorbing or reflecting UV light and thus exhibit variations in floral anthocyanin biosynthesis to scavenge reactive ROS; furthermore, these traits are related to the geographic distribution of cotton species.

Application of Single-Cell Assay for Transposase-Accessible Chromatin with High Throughput Sequencing in Plant Science: Advances, Technical Challenges, and Prospects.

The Single-cell Assay for Transposase-Accessible Chromatin with high throughput sequencing (scATAC-seq) has gained increasing popularity in recent years, allowing for chromatin accessibility to be deciphered and gene regulatory networks (GRNs) to be inferred at single-cell resolution. This cutting-edge technology now enables the genome-wide profiling of chromatin accessibility at the cellular level and the capturing of cell-type-specific cis-regulatory elements (CREs) that are masked by cellular heterogeneity in bulk assays. Additionally, it can also facilitate the identification of rare and new cell types based on differences in chromatin accessibility and the charting of cellular developmental trajectories within lineage-related cell clusters. Due to technical challenges and limitations, the data generated from scATAC-seq exhibit unique features, often characterized by high sparsity and noise, even within the same cell type. To address these challenges, various bioinformatic tools have been developed. Furthermore, the application of scATAC-seq in plant science is still in its infancy, with most research focusing on root tissues and model plant species. In this review, we provide an overview of recent progress in scATAC-seq and its application across various fields. We first conduct scATAC-seq in plant science. Next, we highlight the current challenges of scATAC-seq in plant science and major strategies for cell type annotation. Finally, we outline several future directions to exploit scATAC-seq technologies to address critical challenges in plant science, ranging from plant ENCODE(The Encyclopedia of DNA Elements) project construction to GRN inference, to deepen our understanding of the roles of CREs in plant biology.

Genome-wide identification and expression analysis of the cotton patatin-related phospholipase A genes and response to stress tolerance.

MAIN CONCLUSION: Patatin-related phospholipase A genes were involved in the response of Gossypium hirsutum to drought and salt tolerance. pPLA (patatin-related phospholipase A) is a key enzyme that catalyzes the initial step of lipid hydrolysis, which is involved in biological processes, such as drought, salt stress, and freezing injury. However, a comprehensive analysis of the pPLA gene family in cotton, especially the role of pPLA in the response to drought and salt tolerance, has not been reported so far. A total of 33 pPLA genes were identified in this study using a genome-wide search approach, and phylogenetic analysis classified these genes into three groups. These genes are unevenly distributed on the 26 chromosomes of cotton, and most of them contain a few introns. The results of the collinear analysis showed that G. hirsutum contained 1-5 copies of each pPLA gene found in G. arboreum and G. raimondii. The subcellular localization analysis of Gh_D08G061200 showed that the protein was localized in the nucleus. In addition, analysis of published upland cotton transcriptome data revealed that six GhPLA genes were expressed in various tissues and organs. Two genes (Gh_A04G142100.1 and Gh_D04G181000.1) were highly expressed in all tissues under normal conditions, showing the expression characteristics of housekeeping genes. Under different drought and salt tolerance stresses, we detected four genes with different expression levels. This study helps to clarify the role of pPLA in the response to drought and salt tolerance.

Increasing floral visitation and hybrid seed production mediated by beauty mark in Gossypium hirsutum.

Hybrid crop varieties have been repeatedly demonstrated to produce significantly higher yields than their parental lines; however, the low efficiency and high cost of hybrid seed production has limited the broad exploitation of heterosis for cotton production. One option for increasing the yield of hybrid seed is to improve pollination efficiency by insect pollinators. Here, we report the molecular cloning and characterization of a semidominant gene, Beauty Mark (BM), which controls purple spot formation at the base of flower petals in the cultivated tetraploid cotton species Gossypium barbadense. BM encodes an R2R3 MYB113 transcription factor, and we demonstrate that GbBM directly targets the promoter of four flavonoid biosynthesis genes to positively regulate petal spot development. Introgression of a GbBM allele into G. hirsutum by marker-assisted selection restored petal spot formation, which significantly increased the frequency of honeybee visits in G. hirsutum. Moreover, field tests confirmed that cotton seed yield was significantly improved in a three-line hybrid production system that incorporated the GbBM allele. Our study thus provides a basis for the potentially broad application of this gene in improving the long-standing problem of low seed production in elite cotton hybrid lines.

Integrating advancements in root phenotyping and genome-wide association studies to open the root genetics gateway.

Plant adaptation to challenging environmental conditions around the world has made root growth and development an important research area for plant breeders and scientists. Targeted manipulation of root system architecture (RSA) to increase water and nutrient use efficiency can minimize the adverse effects of climate change on crop production. However, phenotyping of RSA is a major bottleneck since the roots are hidden in the soil. Recently the development of 2- and 3D root imaging techniques combined with the genome-wide association studies (GWASs) have opened up new research tools to identify the genetic basis of RSA. These approaches provide a comprehensive understanding of the RSA, by accelerating the identification and characterization of genes involved in root growth and development. This review summarizes the latest developments in phenotyping techniques and GWAS for RSA, which are used to map important genes regulating various aspects of RSA under varying environmental conditions. Furthermore, we discussed about the state-of-the-art image analysis tools integrated with various phenotyping platforms for investigating and quantifying root traits with the highest phenotypic plasticity in both artificial and natural environments which were used for large scale association mapping studies, leading to the identification of RSA phenotypes and their underlying genetics with the greatest potential for RSA improvement. In addition, challenges in root phenotyping and GWAS are also highlighted, along with future research directions employing machine learning and pan-genomics approaches.

A Rapid and Efficient Method for Isolation and Transformation of Cotton Callus Protoplast.

Protoplasts, which lack cell walls, are ideal research materials for genetic engineering. They are commonly employed in fusion (they can be used for more distant somatic cell fusion to obtain somatic hybrids), genetic transformation, plant regeneration, and other applications. Cotton is grown throughout the world and is the most economically important crop globally. It is therefore critical to study successful extraction and transformation efficiency of cotton protoplasts. In the present study, a cotton callus protoplast extraction method was tested to optimize the ratio of enzymes (cellulase, pectinase, macerozyme R-10, and hemicellulase) used in the procedure. The optimized ratio significantly increased the quantity and activity of protoplasts extracted. We showed that when enzyme concentrations of 1.5% cellulase and 1.5% pectinase, and either 1.5% or 0.5% macerozyme and 0.5% hemicellulase were used, one can obtain increasingly stable protoplasts. We successfully obtained fluorescent protoplasts by transiently expressing fluorescent proteins in the isolated protoplasts. The protoplasts were determined to be suitable for use in further experimental studies. We also studied the influence of plasmid concentration and transformation time on protoplast transformation efficiency. When the plasmid concentration reaches 16 µg and the transformation time is controlled within 12-16 h, the best transformation efficiency can be obtained. In summary, this study presents efficient extraction and transformation techniques for cotton protoplasts.

Computational Drug Repurposing Based on a Recommendation System and Drug-Drug Functional Pathway Similarity.

Drug repurposing identifies new clinical indications for existing drugs. It can be used to overcome common problems associated with cancers, such as heterogeneity and resistance to established therapies, by rapidly adapting known drugs for new treatment. In this study, we utilized a recommendation system learning model to prioritize candidate cancer drugs. We designed a drug-drug pathway functional similarity by integrating multiple genetic and epigenetic alterations such as gene expression, copy number variation (CNV), and DNA methylation. When compared with other similarities, such as SMILES chemical structures and drug targets based on the protein-protein interaction network, our approach provided better interpretable models capturing drug response mechanisms. Furthermore, our approach can achieve comparable accuracy when evaluated with other learning models based on large public datasets (CCLE and GDSC). A case study about the Erlotinib and OSI-906 (Linsitinib) indicated that they have a synergistic effect to reduce the growth rate of tumors, which is an alternative targeted therapy option for patients. Taken together, our computational method characterized drug response from the viewpoint of a multi-omics pathway and systematically predicted candidate cancer drugs with similar therapeutic effects.

Elevation of GhDREB1B transcription by a copy number variant significantly improves chilling tolerance in cotton.

MAIN CONCLUSION: The elevation of transcript levels of GhDREB1B causes the accumulation of osmoregulants and mitigation of reactive oxygen species, which contributes to the enhanced resistance to chilling stress in AiSheng98 cotton. Low temperature is one of the key environmental stresses that impairs cotton growth and restricts fiber productivity. Dehydration responsive element binding (DREB) transcription factors play an important role in cold response in plants by modulating the transcription level of cold-responsive genes to protect the plants from low-temperature stress. Here, we showed that GhDREB1B, a copy number variant in the AiSheng98 (AS98) cotton mutant, significantly improved chilling tolerance in cotton seedlings, while silencing of GhDREB1B made transgenic cotton sensitive to chilling stress in AS98 cotton compared with control plants. Elevated GhDREB1B transcript level activated the expression of major cold-responsive genes. Genome-wide expression profiling by RNA sequencing revealed the upregulation of genes related to fatty acids, lipid proteins, osmoprotection, and anti-oxidative enzymes in AiSheng98. Excessive accumulation of malondialdehyde (MDA) and higher ion leakage rates occurred in wild-type LFH10 plants when compared to those of Aisheng98 during chilling stress, signifying lower chilling tolerance in the wild-type than in Aisheng98. Furthermore, the Aisheng98 mutant under chilling stress accumulated higher levels of free proline and soluble sugar than LFH10 accumulated. These results suggest that GhDREB1B is a positive regulator and its variant can alter the expression patterns of major low-temperature stress-related genes and enhance chilling tolerance in cotton.

A copy number variant at the HPDA-D12 locus confers compact plant architecture in cotton.

Improving yield is a primary mission for cotton (Gossypium hirsutum) breeders; development of cultivars with suitable architecture for high planting density (HPDA) can increase yield per unit area. We characterized a natural cotton mutant, AiSheng98 (AS98), which exhibits shorter height, shorter branch length, and more acute branch angle than wild-type. A copy number variant at the HPDA locus on Chromosome D12 (HPDA-D12), encoding a dehydration-responsive element-binding (DREB) transcription factor, GhDREB1B, strongly affects plant architecture in the AS98 mutant. We found an association between a tandem duplication of a c. 13.5 kb segment in HPDA-D12 and elevated GhDREB1B expression resulting in the AS98 mutant phenotype. GhDREB1B overexpression confers a significant decrease in plant height and branch length, and reduced branch angle. Our results suggest that fine-tuning GhDREB1B expression may be a viable engineering strategy for modification of plant architecture favorable to high planting density in cotton.

Construction of Gossypium barbadense Mutant Library Provides Genetic Resources for Cotton Germplasm Improvement.

Allotetraploid cotton (Gossypium hirsutum and Gossypium barbadense) are cultivated worldwide for its white fiber. For centuries, conventional breeding approaches increase cotton yield at the cost of extensive erosion of natural genetic variability. Sea Island cotton (G. barbadense) is known for its superior fiber quality, but show poor adaptability as compared to Upland cotton. Here, in this study, we use ethylmethanesulfonate (EMS) as a mutagenic agent to induce genome-wide point mutations to improve the current germplasm resources of Sea Island cotton and develop diverse breeding lines with improved adaptability and excellent economic traits. We determined the optimal EMS experimental procedure suitable for construction of cotton mutant library. At M6 generation, mutant library comprised of lines with distinguished phenotypes of the plant architecture, leaf, flower, boll, and fiber. Genome-wide analysis of SNP distribution and density in yellow leaf mutant reflected the better quality of mutant library. Reduced photosynthetic efficiency and transmission electron microscopy of yellow leaf mutants revealed the effect of induced mutations at physiological and cellular level. Our mutant collection will serve as the valuable resource for basic research on cotton functional genomics, as well as cotton breeding.

The kinase domain of CK1δ can be phosphorylated by Chk1.

Members of the casein kinase 1 (CK1) family are key regulators in numerous cellular signal transduction pathways and in order to prevent the development of certain diseases, CK1 kinase activity needs to be tightly regulated. Modulation of kinase activity by site-specific phosphorylation within the C-terminal regulatory domain of CK1δ has already been shown for several cellular kinases. By using biochemical methods, we now identified residues T161, T174, T176, and S181 within the kinase domain of CK1δ as target sites for checkpoint kinase 1 (Chk1). At least residues T176 and S181 show full conservation among CK1δ orthologues from different eukaryotic species. Enzyme kinetic analysis furthermore led to the hypothesis that site-specific phosphorylation within the kinase domain finally contributes to fine-tuning of CK1δ kinase activity. These data provide a basis for the extension of our knowledge about the role of site-specific phosphorylation for regulation of CK1δ and associated signal transduction pathways.

GFF3sort: a novel tool to sort GFF3 files for tabix indexing.

BACKGROUND: The traditional method of visualizing gene annotation data in JBrowse is converting GFF3 files to JSON format, which is time-consuming. The latest version of JBrowse supports rendering sorted GFF3 files indexed by tabix, a novel strategy that is more convenient than the original conversion process. However, current tools available for GFF3 file sorting have some limitations and their sorting results would lead to erroneous rendering in JBrowse. RESULTS: We developed GFF3sort, a script to sort GFF3 files for tabix indexing. Specifically designed for JBrowse rendering, GFF3sort can properly deal with the order of features that have the same chromosome and start position, either by remembering their original orders or by conducting parent-child topology sorting. Based on our test datasets from seven species, GFF3sort produced accurate sorting results with acceptable efficiency compared with currently available tools. CONCLUSIONS: GFF3sort is a novel tool to sort GFF3 files for tabix indexing. We anticipate that GFF3sort will be useful to help with genome annotation data processing and visualization.

CottonFGD: an integrated functional genomics database for cotton.

BACKGROUND: Cotton (Gossypium spp.) is the most important fiber and oil crop in the world. With the emergence of huge -omics data sets, it is essential to have an integrated functional genomics database that allows worldwide users to quickly and easily fetch and visualize genomic information. Currently available cotton-related databases have some weakness in integrating multiple kinds of -omics data from multiple Gossypium species. Therefore, it is necessary to establish an integrated functional genomics database for cotton. DESCRIPTION: We developed CottonFGD (Cotton Functional Genomic Database, https://cottonfgd.org ), an integrated database that includes genomic sequences, gene structural and functional annotations, genetic marker data, transcriptome data, and population genome resequencing data for all four of the sequenced Gossypium species. It consists of three interconnected modules: search, profile, and analysis. These modules make CottonFGD enable both single gene review and batch analysis with multiple kinds of -omics data and multiple species. CottonFGD also includes additional pages for data statistics, bulk data download, and a detailed user manual. CONCLUSION: Equipped with specialized functional modules and modernized visualization tools, and populated with multiple kinds of -omics data, CottonFGD provides a quick and easy-to-use data analysis platform for cotton researchers worldwide.

Co-expression of GR79 EPSPS and GAT yields herbicide-resistant cotton with low glyphosate residues.

Glyphosate-resistant (GR) crops have been adopted on a massive scale by North and South American farmers. Currently, about 80% of the 120 million hectares of the global genetically modified (GM) crops are GR crop varieties. However, the adoption of GR plants in China has not occurred at the same pace, owing to several factors including, among other things, labour markets and the residual effects of glyphosate in transgenic plants. Here, we report the co-expression of codon-optimized forms of GR79 EPSPS and N-acetyltransferase (GAT) genes in cotton. We found five times more resistance to glyphosate with 10-fold reduction in glyphosate residues in two pGR79 EPSPS-pGAT co-expression cotton lines, GGCO2 and GGCO5. The GGCO2 line was used in a hybridization programme to develop new GR cottons. Field trials at five locations during three growing seasons showed that pGR79-pGAT transgenic cotton lines have the same agronomic performance as conventional varieties, but were USD 390-495 cheaper to produce per hectare because of the high cost of conventional weed management practices. Our strategy to pyramid these genes clearly worked and thus offers attractive promise for the engineering and breeding of highly resistant low-glyphosate-residue cotton varieties.

Brassinosteriod Insensitive 2 (BIN2) acts as a downstream effector of the Target of Rapamycin (TOR) signaling pathway to regulate photoautotrophic growth in Arabidopsis.

The components of the target of rapamycin (TOR) signaling pathway have been well characterized in heterotrophic organisms from yeast to humans. However, because of rapamycin insensitivity, embryonic lethality in tor null mutants and a lack of reliable ways of detecting TOR protein kinase in higher plants, the key players upstream and downstream of TOR remain largely unknown in plants. Using engineered rapamycin-sensitive Binding Protein 12-2 (BP12-2) plants, the present study showed that combined treatment with rapamycin and active-site TOR inhibitors (asTORis) results in synergistic inhibition of TOR activity and plant growth in Arabidopsis. Based on this system, we revealed that TOR signaling plays a crucial role in modulating the transition from heterotrophic to photoautotrophic growth in Arabidopsis. Ribosomal protein S6 kinase 2 (S6K2) was identified as a direct downstream target of TOR, and the growth of TOR-suppressed plants could be rescued by up-regulating S6K2. Systems, genetic, and biochemical analyses revealed that Brassinosteriod Insensitive 2 (BIN2) acts as a novel downstream effector of S6K2, and the phosphorylation of BIN2 depends on TOR-S6K2 signaling in Arabidopsis. By combining pharmacological with genetic and biochemical approaches, we determined that the TOR-S6K2-BIN2 signaling pathway plays important roles in regulating the photoautotrophic growth of Arabidopsis.

CK1δ kinase activity is modulated by protein kinase C α (PKCα)-mediated site-specific phosphorylation.

Cellular signal transduction components are usually regulated not only on transcriptional or translational level, but also by posttranslational modifications. Among these, reversible phosphorylation represents the most abundant modification. In general, phosphorylation events are essential for regulating the activity of central signal transduction proteins, also including kinases itself. Members of the CK1 family can be found as central signal transduction proteins in numerous cellular pathways. Due to its wide variety of cellular functions the activity of CK1 family members has to be tightly regulated. We previously reported that PKA and Chk1 are able to phosphorylate CK1δ within its C-terminal regulatory domain, consequently resulting in altered CK1 kinase activity. In the present study, we show by several methods that protein kinase C α (PKCα) as well is able to phosphorylate CK1δ at its C-terminally located residues S328, T329, and S370. Furthermore, we analyze the functional consequences of PKCα-mediated phosphorylation on CK1δ kinase activity. Mutation of S328, T329, or S370 to alanine dramatically alters the kinetic parameters of CK1δ. By using the PKCα-specific inhibitor Go-6983 in a selected cell culture model, we finally show that the in vitro detected regulatory connection between PKCα and CK1δ is also relevant in the cellular context. Taken together, these data contribute to a deeper understanding of cellular signal transduction networks thereby helping to form a basis for the development of future therapeutic concepts.

CK1δ activity is modulated by CDK2/E- and CDK5/p35-mediated phosphorylation.

CK1 protein kinases form a family of serine/threonine kinases which are highly conserved through different species and ubiquitously expressed. CK1 family members can phosphorylate numerous substrates thereby regulating different biological processes including membrane trafficking, cell cycle regulation, circadian rhythm, apoptosis, and signal transduction. Deregulation of CK1 activity and/or expression contributes to the development of neurological diseases and cancer. Therefore, CK1 became an interesting target for drug development and it is relevant to further understand the mechanisms of its regulation. In the present study, Cyclin-dependent kinase 2/Cyclin E (CDK2/E) and Cyclin-dependent kinase 5/p35 (CDK5/p35) were identified as cellular kinases able to modulate CK1δ activity through site-specific phosphorylation of its C-terminal domain. Furthermore, pre-incubation of CK1δ with CDK2/E or CDK5/p35 reduces CK1δ activity in vitro, indicating a functional impact of the interaction between CK1δ and CDK/cyclin complexes. Interestingly, inhibition of Cyclin-dependent kinases by Dinaciclib increases CK1δ activity in pancreatic cancer cells. In summary, these results suggest that CK1δ activity can be modulated by the interplay between CK1δ and CDK2/E or CDK5/p35. These findings extend our knowledge about CK1δ regulation and may be of use for future development of CK1-related therapeutic strategies in the treatment of neurological diseases or cancer.

The role of Somatic embryogenesis receptor-like kinase 1 in controlling pollen production of the Gossypium anther.

In flowering plants, male gametophytes are generated in anthers from microsporocytes. However, more evidence is needed to reveal the genetic mechanisms which regulate the differentiation and interaction of these highly specialized cells in anthers. Here we report the characterization of a series of male-sterile cotton (Gossypium hirsutum) mutants, including mutants with normal fertility, semi-sterility and complete sterility. These mutants are forms of transgenic cotton containing RNAi vectors with partial cDNA fragments of GhSERK1. The GhSERK1 gene encodes a putative leucine-rich repeat receptor protein kinase (LRR-RLK), and generally has 11 domains. In previous research, we found plants containing GhSERK1 produce an abundance of male reproductive tissue. In this paper, three RNAi constructs were designed separately to analyze its function in anther. After the three RNAi vectors were transformed into the cotton, transgenic plants with the specialized fragment exhibited normal fertility or the pollen energy decreased slightly, as ones with the homologous fragments exhibited various degrees of male sterility with different expression levels of GhSERK1 mRNA. In conclusion, for the transgenic plants with conserved fragments, lower expression levels of GhSERK1 mRNA were in transgenic plants, and a higher degree of male sterility was observed. Taking together, these findings demonstrate the GhSERK1 gene has a role in the development of anthers, especially in the formation of pollen grains. Also, we infer there must be another homolog of GhSERK1 in cotton, and both of GhSERK1 and its homolog function redundantly as important control points in controlling anther pollen production.

GhTPPA_2 enhancement of tobacco sugar accumulation and drought tolerance.

Trehalose metabolism plays an important role in plant growth and response to abiotic stress. Trehalose-6-phosphate (Tre6P) can help regulate sugar homeostasis and act as an indication signal for intracellular sugar levels. Crop productivity can be greatly increased by altering the metabolic level of endogenous trehalose in plants, which can optimize the source-sink connection. In this study, the upland cotton GhTPP protein family was first homologously compared and 24 GhTPP genes were found. Transcriptome analysis revealed that GhTPP members had obvious tissue expression specificity. Among them, GhTPPA_2 (Gh_A12G223300.1) was predominantly expressed in leaves and bolls. The results of subcellular localization showed that GhTPPA_2 is localized in the chloroplast. Via PlantCare, we analyzed the promoters and found that the expression of GhTPPA_2 may be induced by light, abiotic stress, and hormones such as abscisic acid, ethylene, salicylic acid and jasmonic acid. In addition, GhTPPA_2 was overexpressed (TPPAoe) in tobacco, and we found that the TPPase activity of TPPAoe tobacco increased by 66 %. Soluble sugar content increased by 39 % and starch content increased by 27 %. Whereas, the transgenic tobacco had obvious growth advantages under 100 mM mannitol stress. Transcriptome sequencing results showed that the differential genes between TPPAoe and control were considerably enriched in functions related to photosynthesis, phosphate group metabolism, and carbohydrate metabolism. This study shows that GhTPPA_2 is involved in regulating sugar metabolism, improving soluble sugar accumulation and drought stress tolerance of tobacco, which provides theoretical basis for research on high yield and drought tolerance of crops.