Cotton Ovules Research Articles

Soil drought during the development of cotton ovule destroyed the antioxidant balance of cotton pistil to hinder the ovule formation

AbstractReproductive failure in cotton caused by drought has been reported to be closely associated with alterations in pistil fertility; however, the mechanism of the effect of drought on pistil fertility in cotton is less studied. We hypothesized that drought would inhibit the ovule formation to alter pistil potential fertility. To address this hypothesis, we conducted a water deficit induction experiment with a cotton cultivar, Dexiamian 1. Results showed that drought damaged the cytological structure of the developing ovules. This resulted in a lower ovule number, finally leading to lower cottonseed number and boll weight. And the decreased ovule number was closely related to the reactive oxygen species (ROS) accumulation in pistil during ovule development. Further analysis of antioxidant metabolism found that in the enzymatic antioxidant system, drought decreased the activities of superoxide dismutase (SOD) and catalase (CAT), resulting in the accumulation of superoxide anion () and hydrogen peroxide (H2O2). Regarding the non‐enzymatic antioxidant system, the elevated glutathione reductase gene (GhGR) expression under drought promoted the glutathione (GSH) accumulation; however, the decreased dehydroascorbate reductase gene (GhDHAR2) expression under drought inhibited the conversion of GSH to ascorbic acid (AsA). Although the increased monodehydroascorbate reductase gene (GhMDHAR) expression under drought promoted AsA accumulation, drought‐induced reduced ascorbate peroxidase gene (GhAPX) expression inhibited the reduction of H2O2 by AsA, which ultimately led to higher AsA content and H2O2 content. We conclude that drought impedes the ovule formation by disturbing pistil's antioxidant metabolic homeostasis to destruct the cytological structure of the developing ovules.

The Effect of a Low Degree of Fluorine Substitution on Cotton Fiber Properties

AbstractCellulose modification often employs chemical processes to tailor its properties and functionalities to fit the demands of a wide range of applications, maximizing its potential as a versatile and sustainable material. From both synthetic and environmental standpoints, one of the ultimate goals is to achieve significant modifications to enhance the end properties of the cellulose while minimizing the number of modified building blocks. The current study demonstrates that a synthetic glucose derivative, 6‐deoxy‐6‐fluoro‐glucose (6F‐Glc), fed into the fertilized cotton ovules, resulted in the accumulation of fluorine inside the cotton fibers with no apparent alteration to their morphology or development. These fibers exhibited a degree of substitution of 0.006, which is 170 times lower than that reported for chemical methods for cellulose modification. However, the physical characterization of the modified fibers showed a surprisingly large impact of this low‐level modification on the cellulose structure (e.g., hydrogen bonding network rearrangement) and a modest increase in the mechanical properties of the fibers. The obtained results exemplify the use of biological systems to introduce low quantities of new functionalities while maximizing the impact on fiber properties.

The transcription factor ERF108 interacts with AUXIN RESPONSE FACTORs to mediate cotton fiber secondary cell wall biosynthesis.

Phytohormones play indispensable roles in plant growth and development. However, the molecular mechanisms underlying phytohormone-mediated regulation of fiber secondary cell wall (SCW) formation in cotton (Gossypium hirsutum) remain largely underexplored. Here, we provide mechanistic evidence for functional interplay between the APETALA2/ethylene response factor (AP2/ERF) transcription factor GhERF108 and auxin response factors GhARF7-1 and GhARF7-2 in dictating the ethylene-auxin signaling crosstalk that regulates fiber SCW biosynthesis. Specifically, in vitro cotton ovule culture revealed that ethylene and auxin promote fiber SCW deposition. GhERF108 RNA interference (RNAi) cotton displayed remarkably reduced cell wall thickness compared with controls. GhERF108 interacted with GhARF7-1 and GhARF7-2 to enhance the activation of the MYB transcription factor gene GhMYBL1 (MYB domain-like protein 1) in fibers. GhARF7-1 and GhARF7-2 respond to auxin signals that promote fiber SCW thickening. GhMYBL1 RNAi and GhARF7-1 and GhARF7-2 virus-induced gene silencing (VIGS) cotton displayed similar defects in fiber SCW formation as GhERF108 RNAi cotton. Moreover, the ethylene and auxin responses were reduced in GhMYBL1 RNAi plants. GhMYBL1 directly binds to the promoters of GhCesA4-1, GhCesA4-2, and GhCesA8-1 and activates their expression to promote cellulose biosynthesis, thereby boosting fiber SCW formation. Collectively, our findings demonstrate that the collaboration between GhERF108 and GhARF7-1 or GhARF7-2 establishes ethylene-auxin signaling crosstalk to activate GhMYBL1, ultimately leading to the activation of fiber SCW biosynthesis.

Genome-wide chromatin accessibility landscape and dynamics of transcription factor networks during ovule and fiber development in cotton

BackgroundThe development of cotton fiber is regulated by the orchestrated binding of regulatory proteins to cis-regulatory elements associated with developmental genes. The cis–trans regulatory dynamics occurred throughout the course of cotton fiber development are elusive. Here we generated genome-wide high-resolution DNase I hypersensitive sites (DHSs) maps to understand the regulatory mechanisms of cotton ovule and fiber development.ResultsWe generated DNase I hypersensitive site (DHS) profiles from cotton ovules at 0 and 3 days post anthesis (DPA) and fibers at 8, 12, 15, and 18 DPA. We obtained a total of 1185 million reads and identified a total of 199,351 DHSs through ~ 30% unique mapping reads. It should be noted that more than half of DNase-seq reads mapped multiple genome locations and were not analyzed in order to achieve a high specificity of peak profile and to avoid bias from repetitive genomic regions. Distinct chromatin accessibilities were observed in the ovules (0 and 3 DPA) compared to the fiber elongation stages (8, 12, 15, and 18 DPA). Besides, the chromatin accessibility during ovules was particularly elevated in genomic regions enriched with transposable elements (TEs) and genes in TE-enriched regions were involved in ovule cell division. We analyzed cis-regulatory modules and revealed the influence of hormones on fiber development from the regulatory divergence of transcription factor (TF) motifs. Finally, we constructed a reliable regulatory network of TFs related to ovule and fiber development based on chromatin accessibility and gene co-expression network. From this network, we discovered a novel TF, WRKY46, which may shape fiber development by regulating the lignin content.ConclusionsOur results not only reveal the contribution of TEs in fiber development, but also predict and validate the TFs related to fiber development, which will benefit the research of cotton fiber molecular breeding.

Cell-specific clock-controlled gene expression program regulates rhythmic fiber cell growth in cotton

BackgroundThe epidermis of cotton ovule produces fibers, the most important natural cellulose source for the global textile industry. However, the molecular mechanism of fiber cell growth is still poorly understood.ResultsHere, we develop an optimized protoplasting method, and integrate single-cell RNA sequencing (scRNA-seq) and single-cell ATAC sequencing (scATAC-seq) to systematically characterize the cells of the outer integument of ovules from wild type and fuzzless/lintless (fl) cotton (Gossypium hirsutum). By jointly analyzing the scRNA-seq data from wildtype and fl, we identify five cell populations including the fiber cell type and construct the development trajectory for fiber lineage cells. Interestingly, by time-course diurnal transcriptomic analysis, we demonstrate that the primary growth of fiber cells is a highly regulated circadian rhythmic process. Moreover, we identify a small peptide GhRALF1 that circadian rhythmically controls fiber growth possibly through oscillating auxin signaling and proton pump activity in the plasma membrane. Combining with scATAC-seq, we further identify two cardinal cis-regulatory elements (CREs, TCP motif, and TCP-like motif) which are bound by the trans factors GhTCP14s to modulate the circadian rhythmic metabolism of mitochondria and protein translation through regulating approximately one third of genes that are highly expressed in fiber cells.ConclusionsWe uncover a fiber-specific circadian clock-controlled gene expression program in regulating fiber growth. This study unprecedentedly reveals a new route to improve fiber traits by engineering the circadian clock of fiber cells.

Comparative proteomic analysis identified proteins and the phenylpropanoid biosynthesis pathway involved in the response to ABA treatment in cotton fiber development

Abscisic acid (ABA) is a plant hormone that plays an important role in cotton fiber development. In this study, the physiological changes and proteomic profiles of cotton (Gossypium hirsutum) ovules were analyzed after 20 days of ABA or ABA inhibitor (ABAI) treatment. The results showed that compared to the control (CK), the fiber length was significantly decreased under ABA treatment and increased under ABAI treatment. Using a tandem mass tags-based quantitative technique, the proteomes of cotton ovules were comprehensively analyzed. A total of 7321 proteins were identified, of which 365 and 69 differentially accumulated proteins (DAPs) were identified in ABA versus CK and ABAI versus CK, respectively. Specifically, 345 and 20 DAPs were up- and down-regulated in the ABA group, and 65 and 4 DAPs were up- and down-regulated in the ABAI group, respectively. The DAPs in the ABA group were mainly enriched in the biosynthesis of secondary metabolites, phenylpropanoid biosynthesis and flavonoid secondary metabolism, whereas the DAPs in the ABAI group were mainly enriched in the indole alkaloid biosynthesis and phenylpropanoid biosynthesis pathways. Moreover, 9 proteins involved in phenylpropanoid biosynthesis were upregulated after ABA treatment, suggesting that this pathway might play important roles in the response to ABA, and 3 auxin-related proteins were upregulated, indicating that auxin might participate in the regulation of fiber development under ABAI treatment.

GhROP6 GTPase modulates auxin accumulation in cotton fibers by regulating cell-specific GhPIN3a localization.

PIN-FORMED- (PIN) mediated polar auxin transport plays a predominant role in most auxin-triggered organogenesis in plants. Global control of PIN polarity at the plasma membrane contributes to the essential establishment of auxin maxima in most multicellular tissues. However, establishment of auxin maxima in single cells is poorly understood. Cotton fibers, derived from ovule epidermal cells by auxin-triggered cell protrusion, provide an ideal model to explore the underlying mechanism. Here, we report that cell-specific degradation of GhPIN3a, which guides the establishment of the auxin gradient in cotton ovule epidermal cells, is associated with the preferential expression of GhROP6 GTPase in fiber cells. In turn, GhROP6 reduces GhPIN3a abundance at the plasma membrane and facilitates intracellular proteolysis of GhPIN3a. Overexpression and activation of GhROP6 promote cell elongation, resulting in a substantial improvement in cotton fiber length.

Single-cell RNA-seq reveals fate determination control of an individual fibre cell initiation in cotton (Gossypium hirsutum).

Cotton fibre is a unicellular seed trichome, and lint fibre initials per seed as a factor determines fibre yield. However, the mechanisms controlling fibre initiation from ovule epidermis are not understood well enough. Here, with single-cell RNA sequencing (scRNA-seq), a total of 14 535 cells were identified from cotton ovule outer integument of Xu142_LF line at four developmental stages (1.5, 1, 0.5 days before anthesis and the day of anthesis). Three major cell types, fibre, non-fibre epidermis and outer pigment layer were identified and then verified by RNA in situ hybridization. A comparative analysis on scRNA-seq data between Xu142 and its fibreless mutant Xu142 fl further confirmed fibre cluster definition. The developmental trajectory of fibre cell was reconstructed, and fibre cell was identified differentiated at 1 day before anthesis. Gene regulatory networks at four stages revealed the spatiotemporal pattern of core transcription factors, and MYB25-like and HOX3 were demonstrated played key roles as commanders in fibre differentiation and tip-biased diffuse growth respectively. A model for early development of a single fibre cell was proposed here, which sheds light on further deciphering mechanism of plant trichome and the improvement of cotton fibre yield.

Development of the engineered “glanded plant and glandless seed” cotton

After fiber, cottonseed is the second most important by-product of cotton production. However, high concentrations of toxic free gossypol deposited in the glands of the cottonseed greatly hamper its effective usage as food or feed. Here, we developed a cotton line with edible cottonseed by specifically silencing the endogenous expression of GoPGF in the seeds, which led to a glandless phenotype with an ultra-low gossypol content in the seeds and nearly normal gossypol in other parts of the plants. This engineered cotton maintains normal resistance to insect pests, but the gossypol content in the seeds dropped by 98%, and thus, it can be consumed directly as food. The trait of a low gossypol content in the cottonseeds was stable and heritable, while the protein, oil content, and fiber yield or quality were nearly unchanged compared to the transgenic receptor W0. In addition, comparative transcriptome analysis showed that down-regulated genes in the ovules of the glandless cotton were enriched in terpenoid biosynthesis, indicating the underlying relationship between gland formation and gossypol biosynthesis. These results pave the way for the comprehensive utilization of cotton as a fiber, oil, and feed crop in the future.

Comparative Metabolomics Analysis Reveals Sterols and Sphingolipids Play a Role in Cotton Fiber Cell Initiation.

Cotton fiber is a seed trichome that protrudes from the outer epidermis of cotton ovule on the day of anthesis (0 day past anthesis, 0 DPA). The initial number and timing of fiber cells are closely related to fiber yield and quality. However, the mechanism underlying fiber initiation is still unclear. Here, we detected and compared the contents and compositions of sphingolipids and sterols in 0 DPA ovules of Xuzhou142 lintless-fuzzless mutants (Xufl) and Xinxiangxiaoji lintless-fuzzless mutants (Xinfl) and upland cotton wild-type Xuzhou142 (XuFL). Nine classes of sphingolipids and sixty-six sphingolipid molecular species were detected in wild-type and mutants. Compared with the wild type, the contents of Sphingosine-1-phosphate (S1P), Sphingosine (Sph), Glucosylceramide (GluCer), and Glycosyl-inositol-phospho-ceramides (GIPC) were decreased in the mutants, while the contents of Ceramide (Cer) were increased. Detail, the contents of two Cer molecular species, d18:1/22:0 and d18:1/24:0, and two Phyto-Cer molecular species, t18:0/22:0 and t18:0/h22:1 were significantly increased, while the contents of all GluCer and GIPC molecular species were decreased. Consistent with this result, the expression levels of seven genes involved in GluCer and GIPC synthesis were decreased in the mutants. Furthermore, exogenous application of a specific inhibitor of GluCer synthase, PDMP (1-phenyl-2-decanoylamino-3-morpholino-1-propanol), in ovule culture system, significantly inhibited the initiation of cotton fiber cells. In addition, five sterols and four sterol esters were detected in wild-type and mutant ovules. Compared with the wild type, the contents of total sterol were not significantly changed. While the contents of stigmasterol and campesterol were significantly increased, the contents of cholesterol were significantly decreased, and the contents of total sterol esters were significantly increased. In particular, the contents of campesterol esters and stigmasterol esters increased significantly in the two mutants. Consistently, the expression levels of some sterol synthase genes and sterol ester synthase genes were also changed in the two mutants. These results suggested that sphingolipids and sterols might have some roles in the initiation of fiber cells. Our results provided a novel insight into the regulatory mechanism of fiber cell initiation.

MiRNA regulation in cotton ovules

miRNA regulation in cotton ovules

A Malvaceae-specific miRNA targeting the newly duplicated GaZIP1L to regulate Zn2+ ion transporter capacity in cotton ovules.

MicroRNAs (miRNAs) play critical roles in regulating gene expression in plants, yet their functions underlying cultivated diploid Gossypium arboreum cotton ovule development are largely unknown. Here, we acquired small RNA profiles from G. arboreum ovules and fibers collected at different growth stages, and identified 46 novel miRNAs that accounted for 23.7% of all miRNAs in G. arboreum reported in the latest plant sRNA database. Through analysis of 84 (including 38 conserved) differentially expressed G. arboreum miRNAs, we detected 215 putative protein-coding genes in 26 biological processes as their potential targets. A Malvaceae-specific novel miRNA named gar-miRN44 was found to likely regulate cotton ovule growth by targeting to a newly duplicated Zn2+ ion transporter gene GaZIP1L. During cotton ovule development, gar-miRN44 transcript level decreased sharply after 10 to 15 days post-anthesis (DPA), while that of the GaZIP1L increased significantly, with a concomitant increase of Zn2+ ion concentration in late ovule developmental stages. Molecular dynamics simulation and ion absorption analysis showed that GaZIP1L has stronger Zn2+ ion binding ability than the original GaZIP1, indicating that the newly evolved GaZIP1L may be more suitable for maintaining high Zn2+ ion transport capacity that is likely required for cotton ovule growth via enhanced cellulose synthase activities. Our systematic miRNA profiling in G. arboreum and characterization of gar-miRN44 not only contribute to the understanding of miRNA function in cotton, but also provide potential targets for plant breeding.

Comparative acetylome analysis of wild-type and fuzzless-lintless mutant ovules of upland cotton (Gossypium hirsutum Cv. Xu142) unveils differential protein acetylation may regulate fiber development

Protein acetylation (KAC) is a significant post-translational modification, which plays an essential role in the regulation of growth and development. Unfortunately, related studies are inadequately available in angiosperms, and to date, there is no report providing insight on the role of protein acetylation in cotton fiber development. Therefore, we first compared the lysine-acetylation proteome (acetylome) of upland cotton ovules in the early fiber development stages by using wild-type as well as its fuzzless-lintless mutant to identify the role of KAC in the fiber development. A total of 1696 proteins with 2754 acetylation sites identified with the different levels of acetylation belonging to separate subcellular compartments suggesting a large number of proteins differentially acetylated in two cotton cultivars. About 80% of the sites were predicted to localize in the cytoplasm, chloroplast, and mitochondria. Seventeen significantly enriched acetylation motifs were identified. Serine and threonine and cysteine located downstream and upstream to KAC sites. KEGG pathway enrichment analysis indicated oxidative phosphorylation, fatty acid, ribosome and protein, and folate biosynthesis pathways enriched significantly. To our knowledge, this is the first report of comparative acetylome analysis to compare the wild-type as well as its fuzzless-lintless mutant acetylome data to identify the differentially acetylated proteins, which may play a significant role in cotton fiber development.

The laccase gene GhLac1 modulates fiber initiation and elongation by coordinating jasmonic acid and flavonoid metabolism

Cotton fibers are single cells originating in the epidermis of cotton ovules, and serve as the largest natural fiber source for the textile industry. In theory, all epidermal cells have the potential to develop into fibers, but only 15%–25% of epidermis cells develop into commercially viable lint fibers. We previously showed that GhLac1 participates in cotton defense against biotic stress. Here we report that GhLac1 also has a role in cotton fiber development. GhLac1 RNAi lines in cotton showed increased differentiation of fiber initials from epidermis and shortened fiber length, resulting in unchanged lint percentage. Suppression of GhLac1 expression led to constitutively hyperaccumulated jasmonic acid (JA) and flavonoids in ovules and fiber cells. In vitro ovule culture experiments confirmed the distinct roles of JA and flavonoids in fiber initiation and elongation, and showed that fiber development is spatially regulated by these chemicals: the increased fiber initiation in GhLac1 RNAi lines is caused by hyperaccumulated JA and rutin content during the fiber initiation stage and shortened fiber length is caused by constitutively increased JA and naringenin content during the fiber elongation stage.

Genome-wide identification and expression analysis of DNA demethylase family in cotton

BackgroundDNA methylation is an important epigenetic factor that maintains and regulates gene expression. The mode and level of DNA methylation depend on the roles of DNA methyltransferase and demethylase, while DNA demethylase plays a key role in the process of DNA demethylation. The results showed that the plant’s DNA demethylase all contained conserved DNA glycosidase domain. This study identified the cotton DNA demethylase gene family and analyzed it using bioinformatics methods to lay the foundation for further study of cotton demethylase gene function.ResultsThis study used genomic information from diploid Gossypium raimondii JGI (D), Gossypium arboreum L. CRI (A), Gossypium hirsutum L. JGI (AD1) and Gossypium barbadebse L. NAU (AD2) to Arabidopsis thaliana. Using DNA demethylase genes sequence of Arabidopsis as reference, 25 DNA demethylase genes were identified in cotton by BLAST analysis. There are 4 genes in the genome D, 5 genes in the genome A, 10 genes in the genome AD1, and 6 genes in the genome AD2. The gene structure and evolution were analyzed by bioinformatics, and the expression patterns of DNA demethylase gene family in Gossypium hirsutum L. were analyzed. From the phylogenetic tree analysis, the DNA demethylase gene family of cotton can be divided into four subfamilies: REPRESSOR of SILENCING 1 (ROS1), DEMETER (DME), DEMETER-LIKE 2 (DML2), and DEMETER-LIKE3 (DML3). The sequence similarity of DNA demethylase genes in the same species was higher, and the genetic relationship was also relatively close. Analysis of the gene structure revealed that the DNA demethylase gene family members of the four subfamilies varied greatly. Among them, the number of introns of ROS1 and DME subfamily was larger, and the gene structure was more complex. For the analysis of the conserved domain, it was known that the DNA demethylase family gene member has an endonuclease III (ENDO3c) domain.ConclusionThe genes of the DNA demethylase family are distributed differently in different cotton species, and the gene structure is very different. High expression of ROS1 genes in cotton were under abiotic stress. The expression levels of ROS1 genes were higher during the formation of cotton ovule. The transcription levels of ROS1 family genes were higher during cotton fiber development.

Association mapping seed kernel oil content in upland cotton using genome-wide SSRs and SNPs

Cotton is a major fiber and oilseed crop; however, few studies have been focused on oil content in cotton. To dissect the genetic basis of seed oil content (SOC), genome-wide association study was performed by using SOC phenotypic data of 503 upland cotton inbred accessions in eight environments and two sets of genotypic data including 179 simple sequence repeats (SSRs) and 11,975 single-nucleotide polymorphism (SNPs) from our previous study. SOC for a given genotype was fairly stable which was reflected in a broad-sense heritability of H2 = 96.6%. Using the mixed linear model (K + Q), 16 significant SSRs (P < 0.01) and 26 significant SNPs (P < 1/11,975), were detected cross BLUP values and at least one individual environments. Based on the LD block analysis, eight SOC-associated QTL on chromosome 6, 10, 12, 13, 15, 17, and 24 covering more than one significant markers were identified, two QTL of them on chromosome 10 and 13 could be matched between SSR and SNP markers. The confidence interval of candidate genes in each QTL ranged from 0.14 to 1.50 Mb, and 12 to 70 genes were identified in these QTL. According to the gene function annotation and gene FPKM value of the 20~35 days post anther (DPA) cotton ovules, key genes involved in lipid metabolism and transport; tricarboxylic acid cycle which might affect seed oil content were found near the leading markers. These results will facilitate the molecular marker assisted breeding of cotton oil content.

Modification of phytosterol composition influences cotton fiber cell elongation and secondary cell wall deposition

BackgroundCotton fiber is a single cell that arises from the epidermis of ovule. It is not only a main economic product of cotton, but an ideal material for studying on the growth and development of plant cell. Our previous study indicated that phytosterol content and the ratio of campesterol to sitosterol fluctuated regularly in cotton fiber development. However, what effects of modified phytosterol content and composition on the growth and development of cotton fiber cell is unknown. In this study, we overexpressed the GhSMT2–1, a cotton homologue of sterol C-24 methyltransferase 2 gene in transgenic upland cotton plants to modify phytosterol content and composition in fiber cells and investigated the changes on fiber elongation and secondary cell wall deposition.ResultsGhSMT2–1 overexpression led to changes of phytosterol content and the ratio of campesterol to sitosterol in fiber cell. At the rapid elongation stage of fiber cell, total phytosterol and sitosterol contents were increased while campesterol content was decreased in transgenic fibers when compared to control fibers. Accordingly, the ratio of campesterol to sitosterol declined strikingly. Simultaneously, the transgenic fibers were shorter and thicker than control fibers. Exogenous application of sitosterol or campesterol separately inhibited control fiber cell elongation in cotton ovule culture system in vitro. In addition, campesterol treatment partially rescued transgenic fiber elongation.ConclusionThese results elucidated that modification of phytosterol content and composition influenced fiber cell elongation and secondary cell wall formation. High sitosterol or low ratio of campesterol to sitosterol suppresses fiber elongation and/or promote secondary cell wall deposition. The roles of sitosterol and campesterol were discussed in fiber cell development. There might be a specific ratio of campesterol to sitosterol in different developmental stage of cotton fibers, in which GhSMT2–1 play an important role. Our study, at a certain degree, provides novel insights into the regulatory mechanisms of fiber cell development.

Identification of Histone H3 (HH3) Genes in Gossypium hirsutum Revealed Diverse Expression During Ovule Development and Stress Responses.

Histone acts as the core for nucleosomes and is a key protein component of chromatin. Among different histone variants, histone H3 (HH3) variants have been reported to play vital roles in plant development. However, biological information and evolutionary relationships of HH3 genes in cotton remain to be elucidated. The current study identified 34 HH3 genes in Gossypium hirsutum. Phylogenetic analysis classified HH3 genes of 19 plant species into eight distinct clades. Sequence logos analysis among Arabidopsis, rice, and G. hirsutum amino acid residues showed higher conservation in amino acids. Using collinearity analysis, we identified 81 orthologous/paralogous gene pairs among the four genomes (A, D, At, and Dt) of cotton. Further, orthologous/paralogous and the Ka/Ks ratio demonstrated that cotton HH3 genes experienced strong purifying selection pressure with restricted functional divergence resulting from segmental and whole genome duplication. Expression pattern analysis indicated that GhHH3 genes were preferentially expressed in cotton ovule tissues. Additionally, GhHH3 gene expression can be regulated by abiotic stresses (cold, heat, sodium chloride (NaCl), and polyethylene glycol (PEG)) and phytohormonal (brassinolide (BL), gibberellic acid (GA), indole-3-acetic acid (IAA), salicylic acid (SA), and methyl jasmonate (MeJA)) treatments, suggesting that GhHH3 genes might play roles in abiotic and hormone stress resistance. Taken together, this work provides important information to decipher complete molecular and physiological functions of HH3 genes in cotton.

Cultures of Gossypium barbadense cotton ovules offer insights into the microtubule-mediated control of fiber cell expansion.

A novel method for culturing ovules of Gossypium barbadense allowed in vitro comparisons with Gossypium hirsutum and revealed variable roles of microtubules in controlling cotton fiber cell expansion. Cotton fibers undergo extensive elongation and secondary wall thickening as they develop into our most important renewable textile material. These single cells elongate at the apex as well as elongating and expanding in diameter behind the apex. These multiple growth modes represent an interesting difference compared to classical tip-growing cells that needs to be explored further. In vitro ovule culture enables experimental analysis of the controls of cotton fiber development in commonly grown Gossypium hirsutum cotton, but, previously, there was no equivalent system for G. barbadense, which produces higher quality cotton fiber. Here, we describe: (a) how to culture the ovules of G. barbadense successfully, and (b) the results of an in vitro experiment comparing the role of microtubules in controlling cell expansion in different zones near the apex of three types of cotton fiber tips. Adding the common herbicide fluridone, 1-Methyl-3-phenyl-5-[3-(trifluoromethyl)phenyl]-4(1H)-pyridinone, to the medium supported G. barbadense ovule culture, with positive impacts on the number of useful ovules and fiber length. The effect is potentially mediated through inhibited synthesis of abscisic acid, which antagonized the positive effects of fluridone. Fiber development was perturbed by adding colchicine, a microtubule antagonist, to ovules of G. barbadense and G. hirsutum cultured 2days after flowering. The results supported the zonal control of cell expansion in one type of G. hirsutum fiber tip and highlighted differences in the role of microtubules in modulating cell expansion between three types of cotton fiber tips.

Cotton fiber elongation requires the transcription factor GhMYB212 to regulate sucrose transportation into expanding fibers.

Cotton is white gold across the globe and composed of fiber cells derived from the outer integument of cotton ovules. Fiber elongation uses sucrose as a direct carbon source. The molecular mechanism transcriptionally controlling sucrose transport from ovules into the elongating fibers remains elusive. In this study the involvement of GhMYB212 in the regulation of sucrose transportion into expanding fibers was investigated. GhMYB212 RNAi plants (GhMYB212i) accumulated less sucrose and glucose in developing fibers, and had shorter fibers and a lower lint index. RNA-seq and protein-DNA binding assays revealed that GhMYB212 was closely linked to the pathways of sucrose and starch transportation and metabolism, directly controling the expression of a sucrose transporter gene GhSWEET12. GhSWEET12 RNAi plants (GhSWEET12i) possessed similar fiber phenotypes to those of GhMYB212i. Exogenous sucrose supplementation in ovule cultures did not rescue the shorter fiber phenotype of GhMYB212i and GhSWEET12i. This finding supported the idea that the attenuated rate of sucrose transport from the outer seed coat into the fibers is responsible for the retardation of fiber elongation. Current investigations support the idea that GhMYB212 functions as the main regulator of fiber elongation by controlling the expression of GhSWEET12, and therefore it is important to study cell expansion and sugar transportation during seed development.