Cotton Fiber Development Research Articles

Brassinosteroids in cotton: orchestrating fiber development

SummaryCotton cultivation spans over 30 million hectares across 85 countries and regions, with more than half participating in the global cotton textile trade. The elongated cotton fiber cell is an ideal model for studying cell elongation and understanding plant growth and development. Brassinosteroids (BRs), recognized for their role in cell elongation, offer the potential for improving cotton fiber quality and yield. Despite extensive research highlighting BR's positive impact on fiber development, a comprehensive review on this topic has been lacking. This review addresses this gap, providing a detailed analysis of the latest advancements in BR signaling and its effects on cotton fiber development. We explore the complex network of BR biosynthesis components, signaling molecules, and regulators, including crosstalk with other pathways and transcriptional control mechanisms. Additionally, we propose molecular strategies and highlight key genetic elements for optimizing BR‐related genes to enhance fiber quality and yield. The review emphasizes the importance of BR homeostasis and the hormonal landscape during cotton fiber development, offering insights into targeted manipulation opportunities and challenges. This consolidation offers a comprehensive understanding of BR's multifaceted roles in fiber development, outlining a strategic approach for BR optimization in cotton fiber quality and yield.

Genetic Regulatory Perturbation of Gene Expression Impacted by Genomic Introgression in Fiber Development of Allotetraploid Cotton.

Interspecific genomic introgression is an important evolutionary process with respect to the generation of novel phenotypic diversity and adaptation. A key question is how gene flow perturbs gene expression networks and regulatory interactions. Here, an introgression population of two species of allopolyploid cotton (Gossypium) to delineate the regulatory perturbations of gene expression regarding fiber development accompanying fiber quality change is utilized. De novo assembly of the recipient parent (G. hirsutum Emian22) genome allowed the identification of genomic variation and introgression segments (ISs) in 323 introgression lines (ILs) from the donor parent (G. barbadense 3-79). It documented gene expression dynamics by sequencing 1,284 transcriptomes of developing fibers and characterized genetic regulatory perturbations mediated by genomic introgression using a multi-locus model. Introgression of individual homoeologous genes exhibiting extreme low or high expression bias can lead to a parallel expression bias in their non-introgressed duplicates, implying a shared yet divergent regulatory fate of duplicated genes following allopolyploidy. Additionally, the IL N182 with improved fiber quality is characterized, and the candidate gene GhFLAP1 related to fiber length is validated. This study outlines a framework for understanding introgression-mediated regulatory perturbations in polyploids, and provides insights for targeted breeding of superior upland cotton fiber.

Cotton Pectate Lyase GhPEL48_Dt Promotes Fiber Initiation Mediated by Histone Acetylation.

GhPEL48_Dt, a Pectate lyase (PEL, EC4.2.2.2), is a crucial enzyme involved in cell-wall modification and pectin degradation. Studies have shown that the GhPEL48_Dt also plays a significant role in cotton-fiber development; however, the specific function and regulatory mechanism of GhPEL48_Dt in cotton-fiber development are still not fully understood. Here, we found that the histone deacetylase inhibitor-Trichostatin A significantly reduces the transcript levels of GhPEL48_Dt and its enzyme activity. Further, silencing of GhPEL48_Dt significantly inhibits the initiation and elongation of cotton fibers by promoting pectin degradation, and the heterologous expression of GhPEL48_Dt promotes the development of trichomes and root hairs in Arabidopsis, which suggests that GhPEL48_Dt plays a positive and conserved role in single cell i.e., fiber, root hair, and leaf trichome development. Collectively, this paper provides a comprehensive analysis of the fundamental characteristics and functions of GhPEL48_Dt in fiber development, including the regulatory role of histone acetylation on GhPEL48_Dt, which contributes to the understanding of pectin degradation pathways and establishes a theoretical foundation for elucidating its regulatory mechanism.

The DUF579 proteins GhIRX15s regulate cotton fiber development by interacting with proteins involved in xylan synthesis

Cotton provides the most abundant natural fiber for the textile industry. The mature cotton fiber largely consists of secondary cell walls with the highest proportion of cellulose and a small amount of hemicellulose and lignin. To dissect the roles of hemicellulosic polysaccharides during fiber development, four IRREGULAR XYLEM 15 (IRX15) genes, GhIRX15-1/-2/-3/-4, were functionally characterized in cotton. These genes encode DUF579 domain-containing proteins, which are homologs of AtIRX15 involved in xylan biosynthesis. The four GhIRX15 genes were predominantly expressed during fiber secondary wall thickening, and the encoded proteins were localized to the Golgi apparatus. Each GhIRX15 gene could restore the xylan deficient phenotype in the Arabidopsis irx15irx15l double mutant. Silencing of GhIRX15s in cotton resulted in shorter mature fibers with a thinner cell wall and reduced cellulose content as compared to the wild type. Intriguingly, GhIRX15-2 and GhIRX15-4 formed homodimers and heterodimers. In addition, the GhIRX15s showed physical interaction with glycosyltransferases GhGT43C, GhGT47A and GhGT47B, which are responsible for synthesis of the xylan backbone and reducing end sequence. Moreover, the GhIRX15s can form heterocomplexes with enzymes involved in xylan modification and side chain synthesis, such as GhGUX1/2, GhGXM1/2 and GhTBL1. These findings suggest that GhIRX15s participate in fiber xylan biosynthesis and modulate fiber development via forming large multiprotein complexes.

GhWRKY40 Interacts with an Asparaginase GhAPD6 Involved in Fiber Development in Upland Cotton (Gossypium hirsutum L.).

Fiber quality improvement is a primary goal in cotton breeding. Identification of fiber quality-related genes and understanding the underlying molecular mechanisms are essential prerequisites. Previously, studies determined that silencing the gene GhWRKY40 resulted in longer cotton fibers; however, both the underlying mechanisms and whether this transcription factor is additionally involved in the regulation of cotton fiber strength/fineness are unknown. In the current study, we verified that GhWRKY40 influences the fiber strength, fiber fineness, and fiber surface structure by using virus-induced gene silencing (VIGS). Potential proteins that may interact with the nucleus-localized GhWRKY40 were screened in a yeast two-hybrid (Y2H) nuclear-system cDNA library constructed from fibers at 0, 10, and 25 days post-anthesis (DPA) in two near-isogenic lines differing in fiber length and strength. An aspartyl protease/asparaginase-related protein, GhAPD6, was identified and confirmed by Y2H and split-luciferase complementation assays. The expression of GhAPD6 was approximately 30-fold higher in the GhWRKY40-VIGS lines at 10 DPA and aspartyl protease activity was significantly upregulated in the GhWRKY40-VIGS lines at 10-20 DPA. This study suggested that GhWRKY40 may interact with GhAPD6 to regulate fiber development in cotton. The results provide a theoretical reference for the selection and breeding of high-quality cotton fibers assisted by molecular technology.

Sphingosine Promotes Fiber Early Elongation in Upland Cotton.

Sphingolipids play an important role in cotton fiber development, but the regulatory mechanism is largely unclear. We found that serine palmitoyltransferase (SPT) enzyme inhibitors, myriocin and sphingosine (dihydrosphingosine (DHS) and phytosphingosine (PHS)), affected early fiber elongation in cotton, and we performed a sphingolipidomic and transcriptomic analysis of control and PHS-treated fibers. Myriocin inhibited fiber elongation, while DHS and PHS promoted it in a dose-effect manner. Using liquid chromatography-tandem mass spectrometry (LC-MS/MS), we found that contents of 22 sphingolipids in the PHS-treated fibers for 10 days were changed, of which the contents of 4 sphingolipids increased and 18 sphingolipids decreased. The transcriptome analysis identified 432 differentially expressed genes (238 up-regulated and 194 down-regulated) in the PHS-treated fibers. Among them, the phenylpropanoid biosynthesis pathway is the most significant enrichment. The expression levels of transcription factors such as MYB, ERF, LBD, and bHLH in the fibers also changed, and most of MYB and ERF were up-regulated. Auxin-related genes IAA, GH3 and BIG GRAIN 1 were up-regulated, while ABPs were down-regulated, and the contents of 3 auxin metabolites were decreased. Our results provide important sphingolipid metabolites and regulatory pathways that influence fiber elongation.

Expression profile analysis of cotton fiber secondary cell wall thickening stage.

To determine the genes associated with the fiber strength trait in cotton, three different cotton cultivars were selected: Sea Island cotton (Xinhai 32, with hyper-long fibers labeled as HL), and upland cotton (17-24, with long fibers labeled as L, and 62-33, with short fibers labeled as S). These cultivars were chosen to assess fiber samples with varying qualities. RNA-seq technology was used to analyze the expression profiles of cotton fibers at the secondary cell wall (SCW) thickening stage (20, 25, and 30 days post-anthesis (DPA)). The results showed that a large number of differentially expressed genes (DEGs) were obtained from the three assessed cotton cultivars at different stages of SCW development. For instance, at 20 DPA, Sea Island cotton (HL) had 6,215 and 5,364 DEGs compared to upland cotton 17-24 (L) and 62-33 (S), respectively. Meanwhile, there were 1,236 DEGs between two upland cotton cultivars, 17-24 (L) and 62-33 (S). Gene Ontology (GO) term enrichment identified 42 functions, including 20 biological processes, 11 cellular components, and 11 molecular functions. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis identified several pathways involved in SCW synthesis and thickening, such as glycolysis/gluconeogenesis, galactose metabolism, propanoate metabolism, biosynthesis of unsaturated fatty acids pathway, valine, leucine and isoleucine degradation, fatty acid elongation pathways, and plant hormone signal transduction. Through the identification of shared DEGs, 46 DEGs were found to exhibit considerable expressional differences at different fiber stages from the three cotton cultivars. These shared DEGs have functions including REDOX enzymes, binding proteins, hydrolases (such as GDSL thioesterase), transferases, metalloproteins (cytochromatin-like genes), kinases, carbohydrates, and transcription factors (MYB and WRKY). Therefore, RT-qPCR was performed to verify the expression levels of nine of the 46 identified DEGs, an approach which demonstrated the reliability of RNA-seq data. Our results provided valuable molecular resources for clarifying the cell biology of SCW biosynthesis during fiber development in cotton.

Exploring the plasmodesmata callose-binding protein gene family in upland cotton: unraveling insights for enhancing fiber length.

Plasmodesmata are transmembrane channels embedded within the cell wall that can facilitate the intercellular communication in plants. Plasmodesmata callose-binding (PDCB) protein that associates with the plasmodesmata contributes to cell wall extension. Given that the elongation of cotton fiber cells correlates with the dynamics of the cell wall, this protein can be related to the cotton fiber elongation. This study sought to identify PDCB family members within the Gossypium. hirsutum genome and to elucidate their expression profiles. A total of 45 distinct family members were observed through the identification and screening processes. The analysis of their physicochemical properties revealed the similarity in the amino acid composition and molecular weight across most members. The phylogenetic analysis facilitated the construction of an evolutionary tree, categorizing these members into five groups mainly distributed on 20 chromosomes. The fine mapping results facilitated a tissue-specific examination of group V, revealing that the expression level of GhPDCB9 peaked five days after flowering. The VIGS experiments resulted in a marked decrease in the gene expression level and a significant reduction in the mature fiber length, averaging a shortening of 1.43-4.77 mm. The results indicated that GhPDCB9 played a pivotal role in the cotton fiber development and served as a candidate for enhancing cotton yield.

Comprehensive Genome-Wide Investigation and Transcriptional Regulation of the DHHC Gene Family in Cotton Seed and Fiber Development

Protein palmitoylation, the most common and the only reversible post-translational lipid modification following protein translation, plays a pivotal role in the biochemical and physiological processes of both animals and plants. DHHC proteins, enriched with DHHC (Asp-His-His-Cys) domains, serve as catalyst for protein palmitoylation. However, research on DHHC in cotton remains scarce. This study conducted a systematic characterization and bioinformatics analysis on G. arboreum, G. raimondii, G. hirsutum, and G. barbadense, detecting 38, 37, 74, and 74 DHHC genes, respectively. Phylogenetic analysis categorized the DHHC gene family into six subgroups, consistent with previous evolutionary studies in Arabidopsis and rice. A further examination of protein structure revealed a correlation between genetic relatedness, structural similarity, and functional identity. Cis-element analysis identified elements predominantly associated with light response, stress, growth and development, and plant hormones. The integration of cotton seed development transcriptome, tissue expression pattern analysis, and population transcriptome data collectively suggests that Ghir_A05G027650 and Ghir_D05G027670 are promising candidate genes influencing seed development in upland cotton. Conversely, Gbar_A04G010750 and Gbar_A12G020520 emerge as potential candidates affecting both seed and fiber development in sea island cotton. These findings lay down a theoretical foundation for delving into the functional diversity of DHHC genes in cotton, thereby paving the way for the development of new breeding strategies and the optimization of cotton seed and fiber production, ultimately contributing to improved crop yield and quality.

Genetic and Molecular Regulation of Cotton Fiber Initiation and Elongation

Cotton fiber, a crucial and sustainable resource for global textile production, undergoes a complex five-stage developmental process, encompassing initiation, elongation, transition, secondary cell wall biosynthesis, and maturation. These elongated single-cell fibers originate from the outer ovule epidermis. The development of cotton fibers involves intricate changes in gene expression and physiological processes, resulting in a nearly pure cellulose product that is vital for the global cotton industry. Decoding the genes associated with fiber development enhances our understanding of cotton fiber mechanisms and facilitates the cultivation of varieties with enhanced quality. In recent decades, advanced omics approaches, including genomics, transcriptomics, and proteomics, have played a pivotal role in identifying the genes and gene products linked to cotton fiber development, including the MYB transcription factor family, which coordinates cotton fiber development. Molecular studies have revealed the transcription factors, like MYB, WRKY, Homeodomain Leucine Zipper (HD-ZIP), and basic helix–loop–helix (bHLH), influencing fiber initiation and elongation. The intricate interplay of phytohormones, like auxin, gibberellic acid (GA), brassinosteroids (BRs), jasmonic acid (JA), ethylene, abscisic acid (ABA), and cytokinin, is explored, providing a comprehensive perspective on the shaping of cotton fibers. Numerous candidate genes and cellular processes affecting various aspects of fiber development hold promise for genetic engineering or marker-assisted breeding to improve fiber quality. This review presents a comprehensive overview of key achievements in cotton molecular biology, with a specific emphasis on recent advancements in understanding the transcription factors and phytohormones involved in cotton fiber initiation and elongation.

Assessment of Segal method for identifying crystallinity evolution in developing cotton fibers

AbstractThe crystallinity index (CI) is an important parameter in evaluating cotton fiber quality. Due to its ease and speed in measuring CIs from X‐ray diffraction (XRD) patterns, the Segal method is popularly used. In this study, we assessed the Segal method for monitoring the crystallinity evolution in developing cotton (Gossypium L.) fibers between 20 and 60 days post anthesis (DPAs) by comparing Segal CIs with those obtained from a Fourier transform infrared spectroscopy‐based method and other XRD‐based methods. The Segal method estimated higher CIs than other methods, especially for shorter DPAs. The Segal method suggested a rapid evolution of crystallinity in the early developmental stage, whereas other methods suggested a gradual increase in crystallinity. The calculation of diffraction patterns for cellulose Iβ crystallites with different sizes showed very little effect of the crystallite size on the Segal CIs for developing cotton fibers studied.

GhATL68b regulates cotton fiber cell development by ubiquitinating the enzyme required for β-oxidation of polyunsaturated fatty acids

E3 ligases are key enzymes required for protein degradation. Here, we identified a C3H2C3 RING domain-containing E3 ubiquitin ligase gene named GhATL68b. It is preferentially and highly expressed in developing cotton fiber cells and shows greater conservation in plants than in animals or archaea. The four orthologous copies of this gene in various diploid cottons and eight in the allotetraploid G. hirsutum were found to have originated from a single common ancestor that can be traced back to Chlamydomonas reinhardtii at about 992 million years ago. Structural variations in the GhATL68b promoter regions of G. hirsutum, G. herbaceum, G. arboreum, and G. raimondii are correlated with significantly different methylation patterns. Homozygous CRISPR-Cas9 knockout cotton lines exhibit significant reductions in fiber quality traits, including upper-half mean length, elongation at break, uniformity, and mature fiber weight. In vitro ubiquitination and cell-free protein degradation assays revealed that GhATL68b modulates the homeostasis of 2,4-dienoyl-CoA reductase, a rate-limiting enzyme for the β-oxidation of polyunsaturated fatty acids (PUFAs), via the ubiquitin proteasome pathway. Fiber cells harvested from these knockout mutants contain significantly lower levels of PUFAs important for production of glycerophospholipids and regulation of plasma membrane fluidity. The fiber growth defects of the mutant can be fully rescued by the addition of linolenic acid (C18:3), the most abundant type of PUFA, to the ovule culture medium. This experimentally characterized C3H2C3 type E3 ubiquitin ligase involved in regulating fiber cell elongation may provide us with a new genetic target for improved cotton lint production.

GhFAD3-4 Promotes Fiber Cell Elongation and Cell Wall Thickness by Increasing PI and IP3 Accumulation in Cotton.

The omega-3 fatty acid desaturase enzyme gene FAD3 is responsible for converting linoleic acid to linolenic acid in plant fatty acid synthesis. Despite limited knowledge of its role in cotton growth, our study focused on GhFAD3-4, a gene within the FAD3 family, which was found to promote fiber elongation and cell wall thickness in cotton. GhFAD3-4 was predominantly expressed in elongating fibers, and its suppression led to shorter fibers with reduced cell wall thickness and phosphoinositide (PI) and inositol triphosphate (IP3) levels. Transcriptome analysis of GhFAD3-4 knock-out mutants revealed significant impacts on genes involved in the phosphoinositol signaling pathway. Experimental evidence demonstrated that GhFAD3-4 positively regulated the expression of the GhBoGH3B and GhPIS genes, influencing cotton fiber development through the inositol signaling pathway. The application of PI and IP6 externally increased fiber length in GhFAD3-4 knock-out plants, while inhibiting PI led to a reduced fiber length in GhFAD3-4 overexpressing plants. These findings suggest that GhFAD3-4 plays a crucial role in enhancing fiber development by promoting PI and IP3 biosynthesis, offering the potential for breeding cotton varieties with superior fiber quality.

Insights into genetic diversity and functional significance of the bHLH genes in cotton fiber development

This study investigated the identification and distribution of single nucleotide polymorphisms (SNPs) within basic Helix-Loop-Helix (bHLH) transcription factors (TFs) across diverse cotton (Gossypium hirsutum) accessions, including wild, landrace, and improved varieties. Comprehensive genomic analyses revealed substantial SNP diversity in wild cotton, distinct from that observed in landraces and improved accessions. Comparative analysis showed notable variations in SNP abundance and impact levels across different accessions, with missense SNPs being prevalent. Nucleotide substitution patterns highlighted the dominance of G>A and A>G substitutions. Population genetic analyses unveiled significant genetic diversity within wild accessions and distinct clustering between wild and improved accessions. Temporal expression profiling of bHLH genes during cotton fiber development demonstrated dynamic expression patterns, emphasizing their roles in fiber initiation, elongation, and cellulose biosynthesis. Moreover, association analysis identified SNPs significantly associated within bHLH genes, particularly GhbHLH149, with fiber quality traits, indicating their potential functional significance. Population genetic analyses further revealed evidence of positive selection on GhbHLH149 during cotton improvement. This study provides a comprehensive understanding of SNP diversity, gene expression dynamics, and genetic selection in bHLH genes, offering valuable insights for future cotton breeding efforts aimed at improving fiber quality traits in cotton.

GhMYB52 Like: A Key Factor That Enhances Lint Yield by Negatively Regulating the Lignin Biosynthesis Pathway in Fibers of Upland Cotton (Gossypium hirsutum L.).

In the context of sustainable agriculture and biomaterial development, understanding and enhancing plant secondary cell wall formation are crucial for improving crop fiber quality and biomass conversion efficiency. This is especially critical for economically important crops like upland cotton (Gossypium hirsutum L.), for which fiber quality and its processing properties are essential. Through comprehensive genome-wide screening and analysis of expression patterns, we identified a particularly high expression of an R2R3 MYB transcription factor, GhMYB52 Like, in the development of the secondary cell wall in cotton fiber cells. Utilizing gene-editing technology to generate a loss-of-function mutant to clarify the role of GhMYB52 Like, we revealed that GhMYB52 Like does not directly contribute to cellulose synthesis in cotton fibers but instead represses a subset of lignin biosynthesis genes, establishing it as a lignin biosynthesis inhibitor. Concurrently, a substantial decrease in the lint index, a critical measure of cotton yield, was noted in parallel with an elevation in lignin levels. This study not only deepens our understanding of the molecular mechanisms underlying cotton fiber development but also offers new perspectives for the molecular improvement of other economically important crops and the enhancement of biomass energy utilization.

Comparative Transcriptomic Analysis of Gossypium hirsutum Fiber Development in Mutant Materials (xin w 139) Provides New Insights into Cotton Fiber Development.

Cotton is the most widely planted fiber crop in the world, and improving cotton fiber quality has long been a research hotspot. The development of cotton fibers is a complex process that includes four consecutive and overlapping stages, and although many studies on cotton fiber development have been reported, most of the studies have been based on cultivars that are promoted in production or based on lines that are used in breeding. Here, we report a phenotypic evaluation of Gossypium hirsutum based on immature fiber mutant (xin w 139) and wild-type (Xin W 139) lines and a comparative transcriptomic study at seven time points during fiber development. The results of the two-year study showed that the fiber length, fiber strength, single-boll weight and lint percentage of xin w 139 were significantly lower than those of Xin W 139, and there were no significant differences in the other traits. Principal component analysis (PCA) and cluster analysis of the RNA-sequencing (RNA-seq) data revealed that these seven time points could be clearly divided into three different groups corresponding to the initiation, elongation and secondary cell wall (SCW) synthesis stages of fiber development, and the differences in fiber development between the two lines were mainly due to developmental differences after twenty days post anthesis (DPA). Differential expression analysis revealed a total of 5131 unique differentially expressed genes (DEGs), including 290 transcription factors (TFs), between the 2 lines. These DEGs were divided into five clusters. Each cluster functional category was annotated based on the KEGG database, and different clusters could describe different stages of fiber development. In addition, we constructed a gene regulatory network by weighted correlation network analysis (WGCNA) and identified 15 key genes that determined the differences in fiber development between the 2 lines. We also screened seven candidate genes related to cotton fiber development through comparative sequence analysis and qRT-PCR; these genes included three TFs (GH_A08G1821 (bHLH), GH_D05G3074 (Dof), and GH_D13G0161 (C3H)). These results provide a theoretical basis for obtaining an in-depth understanding of the molecular mechanism of cotton fiber development and provide new genetic resources for cotton fiber research.

Transgressive and subgenome expression level dominance and co-expression network analyses at the early fiber development in allopolyploid Gossypium

Polyploidization can lead to the emergence of new species. Allotetraploid cotton arose through hybridization and whole genome duplication of its diploid A and D genome progenitors. As a prominent global fiber crop, cotton serves as an excellent model organism for exploring plant evolutionary biology and fiber development, particularly in polyploidization. This study utilized transcriptomics to analyze expression difference between diploid and allotetraploid cotton, aiming to further the understanding of transcriptomic dynamics associated at different fiber developmental stages during polyploidization process. In this study, we identified additive and non-additive genes and clarified the contributions of non-additive genes during the tetraploid formation process. Furthermore, we constructed a gene co-expression network, providing new insights into the transcriptional regulation of polyploid cotton fiber development. Among non-additive genes, more A genome-biased expression level dominance (ELD-A) than D genome (ELD-D) occurred in the tetraploids. In Gossypium hirsutum, ELD-A genes played major roles during initial elongation, exerting greater dominance in fiber development. Transgressive genes impacted early fiber development in Gossypium barbadense. While the At subgenome profoundly influences tetraploid fiber development, Dt sub-genome activation, inheritance and sub/neo-functionalization promote superior high-yield traits. By constructing co-expression networks, we identified key fiber developmental candidate genes, providing valuable resources for functional research and breeding perspectives to develop superior cotton varieties.

Genome-wide identification of the key kinesin genes during fiber and boll development in upland cotton (Gossypium hirsutum L.).

Kinesin is a kind of motor protein, which interacts with microtubule filaments and regulates cellulose synthesis. Cotton fiber is a natural model for studying the cellular development and cellulose synthesis. Therefore, a systematic research of kinesin gene family in cotton (Gossypium spp.) will be beneficial for both understanding the function of kinesin protein and assisting the fiber improvement. Here, we aimed to identify the key kinesin genes present in cotton by combining genome-wide expression profile data, association mapping, and public quantitative trait loci (QTLs) in upland cotton (G. hirsutum L.). Results showed that 159 kinesin genes, including 15 genes of the kinesin-13 gene subfamily, were identified in upland cotton; of which 157 kinesin genes can be traced back to the diploid ancestors, G. raimondii and G. arboreum. Using a combined analysis of public QTLs and genome-wide expression profile information, there were 29 QTLs co-localized together with 28 kinesin genes in upland cotton, including 10 kinesin-13 subfamily genes. Genome-wide expression profile data indicated that, among the 28 co-localized genes, seven kinesin genes were predominantly expressed in fibers or ovules. By association mapping analysis, 30 kinesin genes were significantly associated with three fiber traits, among which a kinesin-13 gene, Ghir_A11G028430, was found to be associated with both cotton boll length and lint weight, and one kinesin-7 gene, Ghir_D04G017880 (Gh_Kinesin7), was significantly associated with fiber strength. In addition, two missense mutations were identified in the motor domain of the Gh_Kinesin7 protein. Overall, the kinesin gene family seemingly plays an important role in cotton fiber and boll development. The exploited kinesin genes will be beneficial for the genetic improvement of fiber quality and yield.

Aspartyl proteases identified as candidate genes of a fiber length QTL, qFLD05, that regulates fiber length in cotton (Gossypium hirsutum L.).

GhAP genes were identified as the candidates involved in cotton fiber length under the scope of fine mapping a stable fiber length QTL, qFLD05. Moreover, the transcription factor GhWRKY40 positively regulated GhAP3 to decrease fiber length. Fiber length (FL) is an economically important fiber quality trait. Although several genes controlling cotton fiber development have been identified, our understanding of this process remains limited. In this study, an FL QTL (qFLD05) was fine-mapped to a 216.9-kb interval using a secondary F2:3 population derived from the upland hybrid cultivar Ji1518. This mapped genomic segment included 15 coding genes, four of which were annotated as aspartyl proteases (GhAP1-GhAP4). GhAPs were identified as candidates for qFLD05 as the sequence variations in GhAPs were associated with FL deviations in the mapping population, and functional validation of GhAP3 and GhAP4 indicated a longer FL following decreases in their expression levels through virus-induced gene silencing (VIGS). Subsequently, the potential involvement of GhWRKY40 in the regulatory network was revealed: GhWRKY40 positively regulated GhAP3's expression according to transcriptional profiling, VIGS, yeast one-hybrid assays and dual-luciferase experiments. Furthermore, alterations in the expression of the eight previously reported cotton FL-responsive genes from the above three VIGS lines (GhAP3, GhAP4 and GhWRKY40) implied that MYB5_A12 was involved in the GhWRKY40-GhAP network. In short, we unveiled the unprecedented FL regulation roles of GhAPs in cotton, which was possibly further regulated by GhWRKY40. These findings will reveal the genetic basis of FL development associated with qFLD05 and be beneficial for the marker-assisted selection of long-staple cotton.

GhHB14_D10 and GhREV_D5, two HD-ZIP III transcription factors, play a regulatory role in cotton fiber secondary cell wall biosynthesis.

GhHB14_D10 and GhREV_D5 regulated secondary cell wall formation and played an important role in fiber development. Cotton serves as an important source of natural fiber, and the biosynthesis of the secondary cell wall plays a pivotal role in determining cotton fiber quality. Nevertheless, the intricacies of this mechanism in cotton fiber remain insufficiently elucidated. This study investigates the functional roles of GhHB14_D10 and GhREV_D5, two HD-ZIP III transcription factors, in secondary cell wall biosynthesis in cotton fibers. Both GhHB14_D10 and GhREV_D5 were found to be localized in the nucleus with transcriptional activation activity. Ectopic overexpression of GhHB14_D10 and GhREV_D5 in Arabidopsis resulted in changed xylem differentiation, secondary cell wall deposition, and expression of genes related to the secondary cell wall. Silencing of GhHB14_D10 and GhREV_D5 in cotton led to enhanced fiber length, reduced cell wall thickness, cellulose contents and expression of secondary cell wall-related genes. Moreover, GhHB14_D10's direct interaction with GhREV_D5, and transcriptional regulation of cellulose biosynthesis genes GhCesA4-4 and GhCesA7-2 revealed their collaborative roles in secondary cell wall during cotton fiber development. Overall, these results shed light on the roles of GhHB14_D10 and GhREV_D5 in secondary cell wall biosynthesis, offering a strategy for the genetic improvement of cotton fiber quality.