Cotton Mutant Research Articles

Cotton Bollworm (H. armigera) Effector PPI5 Targets FKBP17-2 to Inhibit ER Immunity and JA/SA Responses, Enhancing Insect Feeding.

The cotton bollworm causes severe mechanical damage to plants during feeding and leaves oral secretions (OSs) at the mechanical wounds. The role these OSs play in the invasion of plants is still largely unknown. Here, a novel H. armigera effector peptidyl prolyl trans-isomerase 5 (PPI5) was isolated and characterized. PPI5 induces the programmed cell death (PCD) due to the unfolded protein response (UPR) in tobacco leaf. We reveal that PPI5 is important for the growth and development of cotton bollworm on plants, as it renders plants more susceptible to feeding. The GhFKBP17-2, was identified as a host target for PPI5 with peptidyl-prolyl isomerase (PPIase) activity. CRISPR/Cas9 knock-out cotton mutant (CR-GhFKBP17-1/3), VIGS (TRV: GhFKBP17-2) and overexpression lines (OE-GhFKBP17-1/3) were created and the data indicate that GhFKBP17-2 positively regulates endoplasmic reticulum (ER) stress-mediated plant immunity in response to cotton bollworm infestation. We further confirm that PPI5 represses JA and SA levels by downregulating the expression of JA- and SA-associated genes, including JAZ3/9, MYC2/3, JAR4, PR4, LSD1, PAD4, ICS1 and PR1/5. Taken together, our results reveal that PPI5 reduces plant defense responses and makes plants more susceptible to cotton bollworm infection by targeting and suppressing GhFKBP17-2 -mediated plant immunity.

GhEXL3 participates in brassinosteroids regulation of fiber elongation in Gossypium hirsutum.

Cotton fiber (Gossypium hirsutum) serves as an ideal model for investigating the molecular mechanisms of plant cell elongation at the single-cell level. Brassinosteroids (BRs) play a crucial role in regulating plant growth and development. However, the mechanism by which BR influences cotton fiber elongation remains incompletely understood. In this study, we identified EXORDIUM-like (GhEXL3) through transcriptome analysis of fibers from BR-deficient cotton mutant pagoda 1 (pag1) and BRI1-EMS-SUPPRESSOR 1 (GhBES1.4, encoding a central transcription factor of BR signaling) overexpression cotton lines. Knockout of GhEXL3 using CRISPR/Cas9 was found to impede cotton fiber elongation, while its overexpression promoted fiber elongation, suggesting a positive regulatory function for GhEXL3 in fiber elongation. Furthermore, in vitro ovule culture experiments revealed that the overexpression of GhEXL3 partially counteracted the inhibitory effects of brassinazole (BRZ) on cotton fiber elongation, providing additional evidence of GhEXL3 involvement in BR signaling pathways. Moreover, our findings demonstrate that GhBES1.4 directly binds to the E-box (CACGTG) motif in the GhEXL3 promoter region and enhances its transcription. RNA-seq analysis revealed that overexpression of GhEXL3 upregulated the expression of EXPs, XTHs, and other genes associated with fiber cell elongation. Overall, our study contributes to understanding the mechanism by which BR regulates the elongation of cotton fibers through the direct modulation of GhEXL3 expression by GhBES1.4.

Advances in genome sequencing and artificially induced mutation provides new avenues for cotton breeding.

Cotton production faces challenges in fluctuating environmental conditions due to limited genetic variation in cultivated cotton species. To enhance the genetic diversity crucial for this primary fiber crop, it is essential to augment current germplasm resources. High-throughput sequencing has significantly impacted cotton functional genomics, enabling the creation of diverse mutant libraries and the identification of mutant functional genes and new germplasm resources. Artificial mutation, established through physical or chemical methods, stands as a highly efficient strategy to enrich cotton germplasm resources, yielding stable and high-quality raw materials. In this paper, we discuss the good foundation laid by high-throughput sequencing of cotton genome for mutant identification and functional genome, and focus on the construction methods of mutant libraries and diverse sequencing strategies based on mutants. In addition, the important functional genes identified by the cotton mutant library have greatly enriched the germplasm resources and promoted the development of functional genomes. Finally, an innovative strategy for constructing a cotton CRISPR mutant library was proposed, and the possibility of high-throughput screening of cotton mutants based on a UAV phenotyping platform was discussed. The aim of this review was to expand cotton germplasm resources, mine functional genes, and develop adaptable materials in a variety of complex environments.

Integrative Transcriptomics and Proteomics Analysis of a Cotton Mutant yl1 with a Chlorophyll-Reduced Leaf.

Leaf color mutants serve as ideal materials for studying photosynthesis, chlorophyll metabolism, and other physiological processes. Here, we identified a spontaneous yellow-leaf mutant (yl1) with chlorophyll-reduced leaves from G. hirsutum L. cv ZM24. Compare to wild type ZM24 with green leaves, yl1 exhibited patchy yellow leaves and reduced chlorophyll content. To further explore the mechanisms of the patchy yellow phenotype of the mutant plant, the transcriptomics and proteomics profiles were conducted for the mutant and wild types. A total of 9247 differentially expressed genes (DEGs) and 1368 differentially accumulated proteins (DAPs) were identified. Following gene ontology (GO) annotation and KEGG enrichment, the DEGs/DAPs were found to be significantly involved in multiple important pathways, including the obsolete oxidation-reduction process, photosynthesis, light-harvesting, the microtubule-based process, cell redox homeostasis, and the carbohydrate metabolic process. In photosynthesis and the light-harvesting pathway, a total of 39 DAPs/DEGs were identified, including 9 genes in the PSI, 7 genes in the PS II, 9 genes in the light-harvesting chlorophyll protein complex (LHC), 10 genes in the PsbP family, and 4 genes in the cytochrome b6/f complex. To validate the reliability of the omics data, GhPPD1, a DAPs in the PsbP family, was knocked down in cotton using the TRV-based VIGS system, and it was observed that the GhPPD1-silenced plants exhibited patchy yellow color, accompanied by a significant decrease in chlorophyll content. In conclusion, this study integrated transcriptomic and proteomic approaches to gain a deeper understanding of the mechanisms underlying the chlorophyll-reduced leaf phenotype.

GHCU, a Molecular Chaperone, Regulates Leaf Curling by Modulating the Distribution of KNGH1 in Cotton.

Leaf shape is considered to be one of the most significant agronomic traits in crop breeding. However, the molecular basis underlying leaf morphogenesis in cotton is still largely unknown. In this study, through genetic mapping and molecular investigation using a natural cotton mutant cu with leaves curling upward, the causal gene GHCU is successfully identified as the key regulator of leaf flattening. Knockout of GHCU or its homolog in cotton and tobacco using CRISPR results in abnormal leaf shape. It is further discovered that GHCU facilitates the transport of the HD protein KNOTTED1-like (KNGH1) from the adaxial to the abaxial domain. Loss of GHCU function restricts KNGH1 to the adaxial epidermal region, leading to lower auxin response levels in the adaxial boundary compared to the abaxial. This spatial asymmetry in auxin distribution produces the upward-curled leaf phenotype of the cu mutant. By analysis of single-cell RNA sequencing and spatiotemporal transcriptomic data, auxin biosynthesis genes are confirmed to be expressed asymmetrically in the adaxial-abaxial epidermal cells. Overall, these findings suggest that GHCU plays a crucial role in the regulation of leaf flattening through facilitating cell-to-cell trafficking of KNGH1 and hence influencing the auxin response level.

Construction of ethyl methane sulfonate mutant library in G. arboreum and rapid identification of mutant genes via repeated re-sequencing

Cotton (Gossypium) is the leading economic crop and provides most of renewable spinning fibers for the worldwide textile industry. The diploid Asiatic cotton (G. arboreum), with spinnable fiber and relatively simple genome, is an ideal model for cotton functional genomics and fiber biology. In this study, we constructed an ethyl methane sulfonate (EMS) mutant library of Asiatic cotton. Stable mutants with various phenotypic variations in leaf, flower, fiber and plant architecture were obtained in M1 generation. Genome-wide mutation analysis revealed high-density mutations distributed evenly on whole genome. Using repeated re-sequencing of M1 mutant plants, the candidate genes of 10 out of 15 mutants were identified. The functions of GaYUC4 and GaPDX1, candidate genes for bar blade and yellow leaf vein, respectively, were further confirmed using virus induced gene silencing in Asiatic and/or upland cottons. Our work paved way for expansion of Asiatic cotton mutant library and subsequent applications in cotton functional genomics and cotton breeding.

GhSINA1, a SEVEN in ABSENTIA ubiquitin ligase, negatively regulates fiber development in Upland cotton

Protein ubiquitination is essential for plant growth and responses to the environment. The SEVEN IN ABSENTIA (SINA) ubiquitin ligases have been extensively studied in plants, but information on their roles in fiber development is limited. Here, we identified GhSINA1 in Upland cotton (Gossypium hirsutum), which has a conserved RING finger domain and SINA domain. Quantitative real-time PCR (qRT-PCR) analysis showed that GhSINA1 was preferentially expressed during fiber initiation and elongation, especially during initiation in the fuzzless-lintless cotton mutant. Subcellular localization experiments indicated that GhSINA1 localized to the nucleus. In vitro ubiquitination analysis revealed that GhSINA1 has E3 ubiquitin ligase activity. Ectopic overexpression of GhSINA1 in Arabidopsis thaliana reduced the number and length of root hairs and trichomes. Yeast two-hybrid (Y2H), firefly luciferase complementation imaging (LCI), and bimolecular fluorescence complementation (BiFC) assays demonstrated that the GhSINA1 proteins could interact with each other to form homodimers and heterodimers. Overall, these results suggest that GhSINA1 may act as a negative regulator in cotton fiber development through homodimerization and heterodimerization.

Transcriptional and translational landscape fine-tune genome annotation and explores translation control in cotton

IntroductionThe unavailability of intergenic region annotation in whole genome sequencing and pan-genomics hinders efforts to enhance crop improvement. ObjectivesDespite advances in research, the impact of post-transcriptional regulation on fiber development and translatome profiling at different stages of fiber growth in cotton (G. hirsutum) remains unexplored. MethodsWe utilized a combination of reference-guided de novo transcriptome assembly and ribosome profiling techniques to uncover the hidden mechanisms of translational control in eight distinct tissues of upland cotton. ResultsOur study identified P-site distribution at three-nucleotide periodicity and dominant ribosome footprint at 27 nucleotides. Specifically, we have detected 1,589 small open reading frames (sORFs), including 1,376 upstream ORFs (uORFs) and 213 downstream ORFs (dORFs), as well as 552 long non-coding RNAs (lncRNAs) with potential coding functions, which fine-tune the annotation of the cotton genome. Further, we have identified novel genes and lncRNAs with strong translation efficiency (TE), while sORFs were found to affect mRNA transcription levels during fiber elongation. The reliability of these findings was confirmed by the high consistency in correlation and synergetic fold change between RNA-sequencing (RNA-seq) and Ribosome-sequencing (Ribo-seq) analyses. Additionally, integrated omics analysis of the normal fiber ZM24 and short fiber pag1 cotton mutant revealed several differentially expressed genes (DEGs), and fiber-specific expressed (high/low) genes associated with sORFs (uORFs and dORFs). These findings were further supported by the overexpression and knockdown of GhKCS6, a gene associated with sORFs in cotton, and demonstrated the potential regulation of the mechanism governing fiber elongation on both the transcriptional and post-transcriptional levels. ConclusionReference-guided transcriptome assembly and the identification of novel transcripts fine-tune the annotation of the cotton genome and predicted the landscape of fiber development. Our approach provided a high-throughput method, based on multi-omics, for discovering unannotated ORFs, hidden translational control, and complex regulatory mechanisms in crop plants.

A mutant cotton fatty acid desaturase 2-1d allele causes protein mistargeting and altered seed oil composition

BackgroundCotton (Gossypium sp.) has been cultivated for centuries for its spinnable fibers, but its seed oil also possesses untapped economic potential if, improvements could be made to its oleic acid content.ResultsPrevious studies, including those from our laboratory, identified pima accessions containing approximately doubled levels of seed oil oleic acid, compared to standard upland cottonseed oil. Here, the molecular properties of a fatty acid desaturase encoded by a mutant allele identified by genome sequencing in an earlier analysis were analyzed. The mutant sequence is predicted to encode a C-terminally truncated protein lacking nine residues, including a predicted endoplasmic reticulum membrane retrieval motif. We determined that the mutation was caused by a relatively recent movement of a Ty1/copia type retrotransposon that is not found associated with this desaturase gene in other sequenced cotton genomes. The mutant desaturase, along with its repaired isozyme and the wild-type A-subgenome homoeologous protein were expressed in transgenic yeast and stably transformed Arabidopsis plants. All full-length enzymes efficiently converted oleic acid to linoleic acid. The mutant desaturase protein produced only trace amounts of linoleic acid, and only when strongly overexpressed in yeast cells, indicating that the missing C-terminal amino acid residues are not strictly required for enzyme activity, yet are necessary for proper subcellular targeting to the endoplasmic reticulum membrane.ConclusionThese results provide the biochemical underpinning that links a genetic lesion present in a limited group of South American pima cotton accessions and their rare seed oil oleic acid traits. Markers developed to the mutant desaturase allele are currently being used in breeding programs designed to introduce this trait into agronomic upland cotton varieties.

GhBES1 mediates brassinosteroid regulation of leaf size by activating expression of GhEXO2 in cotton (Gossypium hirsutum).

We proposed a working model of BR to promote leaf size through cell expansion. In the BR signaling pathway, GhBES1 affects cotton leaf size by binding to and activating the expression of the E-box element in the GhEXO2 promoter region. Brassinosteroid (BR) is an essential phytohormone that controls plant growth. However, the mechanisms of BR regulation of leaf size remain to be determined. Here, we found that the BR deficient cotton mutant pagoda1 (pag1) had a smaller leaf size than wild-type CRI24. The expression of EXORDIUM (GhEXO2) gene, was significantly downregulated in pag1. Silencing of BRI1-EMS-SUPPRESSOR 1 (GhBES1), inhibited leaf cell expansion and reduced leaf size. Overexpression of GhBES1.4 promoted leaf cell expansion and enlarged leaf size. Expression analysis showed GhEXO2 expression positively correlated with GhBES1 expression. In plants, altered expression of GhEXO2 promoted leaf cell expansion affecting leaf size. Furthermore, GhBES1.4 specifically binds to the E-box elements in the GhEXO2 promoter, inducing its expression. RNA-seq data revealed many down-regulated genes related to cell expansion in GhEXO2 silenced plants. In summary, we discovered a novel mechanism of BR regulation of leaf size through GhBES1 directly activating the expression of GhEXO2.

A 2-bp frameshift deletion at GhDR, which encodes a B-BOX protein that co-segregates with the dwarf-red phenotype in Gossypium hirsutums L.

Plant architecture and leaf color are important factors influencing cotton fiber yield. In this study, based on genetic analysis, stem paraffin sectioning, and phytohormone treatments, we showed that the dwarf-red (DR) cotton mutant is a gibberellin-sensitive mutant caused by a mutation in a single dominant locus, designated GhDR. Using bulked segregant analysis (BSA) and genotyping by target sequencing (GBTS) approaches, we located the causative mutation to a ~197-kb genetic interval on chromosome A09 containing 25 annotated genes. Based on gene annotation and expression changes between the mutant and normal plants, GH_A09G2280 was considered to be the best candidate gene responsible for the dwarf and red mutant phenotypes. A 2-nucleotide deletion was found in the coding region of GhDR/GH_A09G2280 in the DR mutant, which caused a frameshift and truncation of GhDR. GhDR is a homolog of Arabidopsis AtBBX24, and encodes a B-box zinc finger protein. The frameshift deletion eliminated the C-terminal nuclear localization domain and the VP domain of GhDR, and altered its subcellular localization. A comparative transcriptome analysis demonstrated downregulation of the key genes involved in gibberellin biosynthesis and the signaling transduction network, as well as upregulation of the genes related to gibberellin degradation and the anthocyanin biosynthetic pathway in the DR mutant. The results of this study revealed the potential molecular basis by which plant architecture and anthocyanin accumulation are regulated in cotton.

EVALUATION OF COTTON MUTANTS FOR WATER DEFICIT CONDITION

Cotton (Gossypium hirsutum ssp.) belongs to the genus Gossypium from the family Malvaceae. About 100 genera are in the family Malvaceae and about 1500 species. It is grown worldwide for oil, seed, and fiber. A major problem in cotton cultivation is a drought that’s why this experiment was planned to evaluate the cotton mutants for drought tolerance. A variety of cotton Cyto-155 was mutated with gamma rays at different intensities such that 20kR, 25kR, 30kR, and 35kR, to obtain some genetic variation. Mutated seed along with non-mutated of that cotton variety was sown in split plot layout under randomized complete block design with three replications, under the normal irrigated condition with six irrigations and water stress condition with two irrigations at maturity data was collected for plant height, number of monopodial branches, number of sympodial branches, bolls per plant, boll weight, seed index, GOT%, lint index, seed cotton yield, relative water content and excised leaf water loss.Results showed a highly significant effect of genotype and water regime on all observed traits. Water regime × genotype interaction was also highly significant for plant height, number of sympodial branches, seed index, boll weight, lint index, and excised leaf water loss. Water regime × genotype interaction was non-significant for number of monopodial branches, bolls per plant, GOT%, seed cotton yield, and relative water content. Mean comparison indicated that seed mutated at 30kR and 35kR performed well for all yield-related traits in both irrigated conditions.

Deficiency of a peroxisomal NADP-isocitrate dehydrogenase leads to dwarf plant and defect seed in upland cotton.

The NADP-isocitrate dehydrogenase-encoded gene GH_D13G1452 with a C-terminus tripeptide Proline-Lysine-Leucine was localized in the peroxisome. It was highly expressed in stems and ovules of 15 days post-anthesis and responded to multiple external stimuli in upland cotton. An upland cotton mutant (Ghpericdh) was identified by flanking sequence amplification and genome variation detection that exogenous sequence was inserted in the middle of the 12th intron of GH_D13G1452, resulting in the deficiency of gene expression. The Ghpericdh mutant displayed a dwarf plant phenotype when grown under field or greenhouse conditions, and GH_D13G1452 functioned as an incomplete dominance on plant height. The germination rate of mutant seed from greenhouse-grown plants was dramatically lower than that from field-grown plants, which indicated that GhperICDH plays a critical role in seed maturation and germination. Therefore, GH_D13G1452 is indispensable in the development of stems and seeds and functions in the adaptability of cotton to the environment. The Ghpericdh mutant provides insight into the function of peroxisomal ICDH and may contribute to the genetic improvement in cotton.

Isolation and characterization of a new imidazolinone‐tolerant mutant in cotton

AbstractCotton (Gossypium hirsutum L.) cultivars resistant to different herbicides have generated important advances in crop management. There is a wide range of cotton resistance to herbicides, however, commercial imidazolinone (IMI)‐tolerant cultivars are not available. In this research, a new mutation that confers tolerance to IMI in cotton was isolated in the M2 generation after seed mutagenic treatments with sodium azide water solutions. The mode of inheritance of the trait was evaluated in the F2 generation by the application of imazethapyr. The inheritance corresponds to a semidominant type with a 1:2:1 distribution of tolerant/intermediate tolerance/not tolerant, in agreement with that observed in most IMI‐tolerant crops. The IMI‐tolerant line behavior was evaluated in the field nursery after the application of the herbicide in the form of foliar and pre‐emergence sprays. The tolerance level was high in pre‐emergence applications and intermediate in foliar applications compared with the nontolerant genotype, which showed a decrease in fiber yield and quality with both methods of application. After sequencing the acetohydroxyacid synthase (AHAS) gene, a point mutation corresponding to Ala205Val in the AHAS enzyme was found, which suggests it could be responsible for the increased IMI tolerance observed in cotton line SP 4172‐32. Based on results from field assays, it can be concluded that this mutant is a promising experimental line that can be used in obtaining IMI‐ tolerant commercial cotton cultivars.

A deletion/duplication in the Ligon lintless-2 locus induces siRNAs that inhibit cotton fiber cell elongation.

Most cultivated cotton (Gossypium hirsutum L.) varieties have two types of seed fibers: short fuzz fiber strongly adhered to the seed coat, and long lint fiber used in the textile industry. The Ligon lintless-2 (Li2) cotton mutant has a normal vegetative phenotype but produces very short lint fiber on the seeds. The Li2 mutation is controlled by a single dominant gene. We discovered a large structural rearrangement at the end of chromosome D13 in the Li2 mutant based on whole-genome sequencing and genetic mapping of segregating populations. The rearrangement contains a 177-kb deletion and a 221-kb duplication positioned as a tandem inverted repeat. The gene Gh_D13G2437 is located at the junction of the inverted repeat in the duplicated region. During transcription such structure spontaneously forms self-complementary hairpin RNA of Gh_D13G2437 followed by production of small interfering RNA (siRNA). Gh_D13G2437 encodes a Ran-Binding Protein 1 (RanBP1) that preferentially expresses during cotton fiber elongation. The abundance of siRNA produced from Gh_D13G2437 reciprocally corresponds with the abundance of highly homologous (68%-98% amino acid sequence identity) RanBP1 family transcripts during fiber elongation, resulting in a shorter fiber phenotype in the Li2. Overexpression of Gh_D13G2437 in the Li2 mutant recovered the long lint fiber phenotype. Taken together, our findings revealed that siRNA-induced silencing of a family of RanBP1s inhibit elongation of cotton fiber cells in the Li2 mutant.

Loss of function of VdDrs2, a P4-ATPase, impairs the toxin secretion and microsclerotia formation, and decreases the pathogenicity of Verticillium dahliae

Four P4-ATPase flippase genes, VdDrs2, VdNeo1, VdP4-4, and VdDnf1 were identified in Verticillium dahliae, one of the most devastating phytopathogenic fungi in the world. Knock out of VdDrs2, VdNeo1, and VdP4-4, or knock down of VdDnf1 significantly decreased the pathogenicity of the mutants in cotton. Among the mutants, the greatest decrease in pathogenicity was observed in ΔVdDrs2. VdDrs2 was localized to plasma membrane, vacuoles, and trans-Golgi network (TGN). In vivo observation showed that the infection of the cotton by ΔVdDrs2 was significantly delayed. The amount of two known Verticillium toxins, sulfacetamide, and fumonisin B1 in the fermentation broth produced by the ΔVdDrs2 strain was significantly reduced, and the toxicity of the crude Verticillium wilt toxins to cotton cells was attenuated. In addition, the defect of VdDrs2 impaired the synthesis of melanin and the formation of microsclerotia, and decreased the sporulation of V. dahliae. Our data indicate a key role of P4 ATPases-associated vesicle transport in toxin secretion of disease fungi and support the importance of mycotoxins in the pathogenicity of V. dahliae.

Creation of cotton mutant library based on linear electron accelerator radiation mutation

Cotton (Gossypium spp.) is one of the most important cash crops worldwide. At present, new cotton varieties are mainly produced through conventional cross breeding, which is limited by available germplasm. Although the genome of cotton has been fully sequenced, research on the function of specific genes lags behind due to the lack of sufficient genetic material. Therefore, it is very important to create a cotton mutant library to create new, higher-quality varieties and identify genes associated with the regulation of key traits. Traditional mutagenic strategies, such as physical, chemical, and site-directed mutagenesis, are relatively costly, inefficient, and difficult to perform. In this study, we used a radiation mutation method based on linear electron acceleration to mutate cotton variety ‘TM-1’, for which a whole-genome sequence has previously been performed, to create a high throughput cotton mutant library. Abundant phenotypic variation was observed in the progeny population for three consecutive generations, including cotton fiber color variation, plant dwarfing, significant improvement of yield traits, and increased sensitivity to Verticillium wilt. These results show that radiation mutagenesis is an effective and feasible method to create plant mutant libraries.

RNA-Seq with a novel glabrous-ZM24fl reveals some key lncRNAs and the associated targets in fiber initiation of cotton

BackgroundCotton fiber is an important natural resource for textile industry and an excellent model for cell biology study. Application of glabrous mutant cotton and high-throughput sequencing facilitates the identification of key genes and pathways for fiber development and cell differentiation and elongation. LncRNA is a type of ncRNA with more than 200 nt in length and functions in the ways of chromatin modification, transcriptional and post-transcriptional modification, and so on. However, the detailed lncRNA and associated mechanisms for fiber initiation are still unclear in cotton.ResultsIn this study, we used a novel glabrous mutant ZM24fl, which is endowed with higher somatic embryogenesis, and functions as an ideal receptor for cotton genetic transformation. Combined with the high-throughput sequencing, fatty acid pathway and some transcription factors such as MYB, ERF and bHLH families were identified the important roles in fiber initiation; furthermore, 3,288 lncRNAs were identified, and some differentially expressed lncRNAs were also analyzed. From the comparisons of ZM24_0 DPA vs ZM24_-2 DPA and fl_0 DPA vs ZM24_0 DPA, one common lncRNA MSTRG 2723.1 was found that function upstream of fatty acid metabolism, MBY25-mediating pathway, and pectin metabolism to regulate fiber initiation. In addition, other lncRNAs MSTRG 3390.1, MSTRG 48719.1, and MSTRG 31176.1 were also showed potential important roles in fiber development; and the co-expression analysis between lncRNAs and targets showed the distinct models of different lncRNAs and complicated interaction between lncRNAs in fiber development of cotton.ConclusionsFrom the above results, a key lncRNA MSTRG 2723.1 was identified that might mediate some key genes transcription of fatty acid metabolism, MYB25-mediating pathway, and pectin metabolism to regulate fiber initiation of ZM24 cultivar. Co-expression analysis implied that some other important lncRNAs (e.g., MSTRG 3390.1, MSTRG 48719.1, and MSTRG 31176.1) were also showed the different regulatory model and interaction between them, which proposes some valuable clues for the lncRNAs associated mechanisms in fiber development.

Glucose regulates cotton fiber elongation by interacting with brassinosteroid.

In plants, glucose (Glc) plays important roles, as a nutrient and signal molecule, in the regulation of growth and development. However, the function of Glc in fiber development of upland cotton (Gossypium hirsutum) is unclear. Here, using gas chromatography-mass spectrometry (GC-MS), we found that the Glc content in fibers was higher than that in ovules during the fiber elongation stage. In vitro ovule culture revealed that lower Glc concentrations promoted cotton fiber elongation, while higher concentrations had inhibitory effects. The hexokinase inhibitor N-acetylglucosamine (NAG) inhibited cotton fiber elongation in the cultured ovules, indicating that Glc-mediated fiber elongation depends on the Glc signal transduced by hexokinase. RNA sequencing (RNA-seq) analysis and hormone content detection showed that 150mM Glc significantly activated brassinosteroid (BR) biosynthesis, and the expression of signaling-related genes was also increased, which promoted fiber elongation. In vitro ovule culture clarified that BR induced cotton fiber elongation in a dose-dependent manner. In hormone recovery experiments, only BR compensated for the inhibitory effects of NAG on fiber elongation in a Glc-containing medium. However, the ovules cultured with the BR biosynthetic inhibitor brassinazole and from the BR-deficient cotton mutant pag1 had greatly reduced fiber elongation at all the Glc concentrations tested. This demonstrates that Glc does not compensate for the inhibition of fiber elongation caused by BR biosynthetic defects, suggesting that the BR signaling pathway works downstream of Glc during cotton fiber elongation. Altogether, our study showed that Glc plays an important role in cotton fibre elongation, and crosstalk occurs between Glc and BR signaling during modulation of fiber elongation.

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

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.