Shading treatment during late stage of seed development promotes subsequent seed germination and seedlings establishment in sunflower.

During the sunflower seed production process, the role of artificial shading treatment (ST) in seed development and subsequent seed germination remains largely unknown. In the present study, sunflower mother plants were artificially shaded during 1-34 (full period-ST, FST), 1-22 (early period-ST, EST), and 22-34 (late period-ST, LST) days after pollination (DAP), to examine the effects of parental shading on subsequent seed germination. Both FST and EST significantly reduced the photosynthetic efficiency of sunflower, manifested as decreased seed dry weight and unfavorable seed germination. On the contrary, LST remarkably increased seed dry weight and promoted subsequent seed germination and seedling establishment. LST enhanced the activities of several key enzymes involved in triglyceride anabolism and corresponding-genes expression, which in turn increased the total fatty acid contents and altered the fatty acid composition. During early germination, the key enzyme activities involved in triglyceride disintegration and corresponding-gene expressions in LST seeds were apparently higher than those in seeds without the shading treatment (WST). Consistently, LST seeds had significant higher contents of ATP and soluble sugar. Moreover, enzyme activities related to abscisic acid (ABA) biosynthesis and corresponding gene expressions decreased within LST seeds, whereas the enzyme activities and corresponding gene expressions associated with gibberellin (GA) biosynthesis were increased. These results were also evidenced by the reduced ABA content but elevated GA level within LST seeds, giving rise to higher GA/ABA ratio. Our findings suggested that LST could promote sunflower seed development and subsequent seed germination as well as seedling establishment through modulating the dynamic metabolism of triglycerides, fatty acid and GA/ABA balance.

Gossypium purpurascens genome provides insight into the origin and domestication of upland cotton.

INTRODUCTION: Allotetraploid upland cotton (Gossypium hirsutum L.) is native to the Mesoamerican and Caribbean regions, had been improved in the southern United States by the mid-eighteenth century, was then dispersed worldwide. However, a Hainan Island Native Cotton (HIC) has long been grown extensively on Hainan Island, China. OBJECTIVES: Explore HIC's evolutionary relationship and genomic diversity with other tetraploid cottons, its origin and whether it was used for YAZHOUBU (Yazhou cloth, World Intangible Cultural Heritage) weaving, and the role of structural variations (SVs) in upland cotton domestication. METHODS: We assembled a high-quality genome of one HIC plant. We performed phylogenetic analysis, divergence time estimation, principal component analysis and population differentiation estimation using cotton assemblies and/or resequencing data. SVs were detected by whole-genome comparison. A F2 population was used for linkage analysis and to study effects of SVs. Buoyancy and salt water tolerance tests for seeds were conducted. RESULTS: We found that the HIC belongs to G. purpurascens. G. purpurascens is best classified as a primitive race of G. hirsutum. The potential for long range transoceanic dispersal of G. purpurascens seeds was proved. A set of SVs, selective sweep regions between G. hirsutum races and cultivars, and quantitative trait loci (QTLs) of eleven agronomic traits were obtained. SVs, especially large-scale SVs, were found to have important effects on cotton domestication and improvement. Of them, eight large-scale inversions strongly associated with yield and fiber quality have probably undergone artificial selection in domestication. CONCLUSION: G. purpurascens including HIC is a primitive race of G. hirsutum, probably disperse to Hainan from Central America by floating on ocean currents, may have been partly domesticated, planted and was likely used for YAZHOUBU weaving in Hainan much earlier than the Pre-Columbian period. SV plays an important role in cotton domestication and improvement.

Spermidine enhances heat tolerance of rice seeds during mid-filling stage and promote subsequent seed germination.

Introduction: Heat stress is a vital factor which restricts rice seed quality and yield. However, the response mechanism to heat stress in the mid filling stage of rice seed is unclear. Methods: In the present study we integrated phenotypic analysis with biochemical, hormone, and gene expression analysis in order to explore technologies for improving rice seeds heat tolerance and subsequent seed germination. Results: Spermidine (Spd) application effectively alleviated the damage of heat stress treatment during mid-filling stage (HTM, 12-20 days after pollination) on seed development, promoted subsequent seed germination and seedlings establishment. Spd significantly increased seed dry weight, starch and amylose contents during seed development under heat stress, and improved seed germinate, seedlings establishment and seedling characteristics during germination time. Biochemical analysis indicated that, HTM significantly decreased the activities of several starch synthase enzymes and led to a decrease in starch content. While Spd treatment significantly enhanced the activities of ADP-glucose pyrophosphorylas and granule-bound starch synthase, as well as the corresponding-genes expressions in HTM rice seeds, resulting in the increases of amylose and total starch contents. In addition, Spd significantly increased the catalase and glutathione reductase activities together with corresponding-genes expressions, and lowered the overaccumulation of H2O2 and malondialdehyde in HTM seeds. In the subsequent seed germination process, HTM+Spd seeds exhibited dramatically up-regulated levels of soluble sugars, glucose, ATP and energy charges. Consistently, HTM+Spd seeds showed significantly increased of α-amylose and α-glucosidase activities as well as corresponding-genes expressions during early germination. Moreover, HTM evidently increased the abscisic acid (ABA) content, decreased the gibberellin (GA) content, and accordingly significantly declined the GA/ABA ratio during early rice seeds germination. However, Spd treatment did not significantly affect the metabolism of GA and ABA in seed germination stage. Discussion: The present study suggested that Spd treatment could effectively alleviate the negative impact of HTM on seed development and the subsequent seed germination, which might be closely correlated with starch synthesis and antioxidant defense during seed filling period, starch decomposition and energy supply in seed germination period.

Spermidine enhances chilling tolerance of kale seeds by modulating ROS and phytohormone metabolism.

Chilling stress is an important constraint for kale seed germination and seedlings establishment. It is vital to develop an effective approach to enhance kale seed germination ability under chilling stress. The present study reported that spermidine (Spd) could improve seed chilling tolerance in two kale cultivars 'Nagoya' (MGW) and 'Pigeon' (BB) during germination. The results showed that MGW was cold tolerant with a 90.67% germination percentage (GP) under chilling stress, while BB was cold sensitive with a 70.67% GP under chilling stress. Spd content in MGW and BB seeds during seed germination were up-regulated and down-regulated by chilling stress, respectively. Besides, chilling stress apparently decreased the gibberellin (GA) and ethylene (ET) contents, while increased the levels of abscisic acid (ABA) and reactive oxygen species (ROS) in MGW and BB seeds during germination. Exogenous Spd application increased GA, ET contents and decreased ABA content through regulating the gene expressions of metabolic-related enzymes, thus effectively alleviating the low temperature damage on kale seed germination. Besides, Spd significantly increased the activities of superoxide dismutase (SOD) and peroxidase (POD), and reduced the levels of hydrogen peroxide (H2O2) and superoxide anion (O2·-). The present study demonstrated that endogenous Spd metabolism plays an important role in kale seed germination under chilling stress. The effect of exogenous Spd on the metabolism of endogenous Spd, GA, ABA, ET and antioxidant enzymes might be the important reason for promoting the kale seed vigor at low temperature.

Classification and Expression Profile of the U-Box E3 Ubiquitin Ligase Enzyme Gene Family in Maize (Zea mays L.).

The U-box E3 (PUB) family genes encode the E3 ubiquitin ligase enzyme, which determines substrate specific recognition during protein ubiquitination. They are widespread in plants and are critical for plant growth, development, and response to external stresses. However, there are few studies on the functional characteristic of PUB gene family in the important staple crop, maize (Zea mays L.). In this study, the PUB gene in maize was aimed to identify and classify through whole-genome screening. Phylogenetic tree, gene structure, conserved motif, chromosome location, gene duplication (GD), synteny, and cis-acting regulatory element of PUB member were analyzed. The expression profiles of ZmPUB gene family in maize during development and under abiotic stress and hormones treatment were analyzed by the RNA-seq data. A total of 79 PUB genes were identified in maize genome, and they were stratified into seven categories. There were 25 pairs of segmental duplications (SD) and 1 pair of tandem duplication (TD) identified in the maize PUB gene family. A close relationship was observed between the monocot plant maize and rice in PUB gene family. There were 94 kinds of cis-acting elements identified in the maize PUB gene family, which included 46 biotic- and abiotic-responsive elements, 19 hormone-responsive elements, 13 metabolic and growth-related elements. The expression profiles of maize PUB gene family showed characteristics of tissue specificity and response to abiotic stress and hormones treatment. These results provided an extensive overview of the maize PUB gene family.

Ultrasonic Waves Regulate Antioxidant Defense and Gluconeogenesis to Improve Germination From Naturally Aged Soybean Seeds.

Soybean seeds contain substantial triacylglycerols and fatty acids that are prone to oxidation during storage, contributing to the dramatic deterioration of seed vigor. This study reports an ultrasonic waves treatment (UWT), which is a physical method capable of promoting the germination ability of the aged soybean seeds by regulating the antioxidant defense and gluconeogenesis. Germination test revealed that UWT significantly increased the germination rate and seedlings' establishment of the soybean seeds stored for 12 months, although insignificantly impacting the vigor of fresh (stored for 1 month) and short-term stored (for 6 months) seeds. Further biochemical analysis revealed that UWT decreased the hydrogen peroxide (H2O2), O2⋅-, and malondialdehyde contents in the aged soybean seeds during early germination. Consistently, UWT prominently elevated the activities of superoxide dismutase, catalase, and acetaldehyde dehydrogenase, and also the corresponding gene expressions. Besides, the soluble sugar content of UWT was significantly higher than that of the untreated aged seeds. Analysis of enzyme activity showed UWT significantly upregulated the activities of several key enzymes in gluconeogenesis and the transcription levels of corresponding genes. Moreover, UWT enhanced the invertase activity within aged seeds, which was responsible for catalyzing sucrose hydrolysis for forming glucose and fructose. In summary, UWT improved germination and seedlings establishment of aged soybean seeds by regulating antioxidant defense and gluconeogenesis. This study expands the application of ultrasonication in agricultural production and further clarifies the physiological and molecular mechanisms of the aged seed germination, aiming to provide theoretical and practical guidance for seed quality and safety.

Role of phasiRNAs from two distinct phasing frames of GhMYB2 loci in cis- gene regulation in the cotton genome.

BACKGROUND: Phased small interfering RNA (phasiRNA) is primarily derived from the 22-nt miRNA targeting loci. GhMYB2, a gene with potential roles in cotton fiber cell fate determination, is a target gene of miR828 and miR858 in the generation of phasiRNAs. RESULTS: In the presented work, through the evaluation of phasing scores and phasiRNA distribution pattern, we found that phasiRNAs from GhMYB2 were derived from the 3' cleavage fragments of 22-nt miR828 and 21-nt miR858 respectively. These two miRNA targeting sites initiated two phasing frames on transcripts of one locus. By means of RNA ligase-mediated rapid amplification of cDNA ends (RLM-RACE), we further demonstrated that phasiRNAs derived from the two phasing frames played a role in cis-regulation of GhMYB2. The phasiRNAs derived from GhMYB2 were expressed in the somatic tissues, especially in anther and hypocotyl. We further employed our previous small RNA sequencing data as well as the degradome data of cotton fiber bearing ovules, anthers, hypocotyls and embryogenic calli tissues published in public databases, to validate the expression, phasing pattern and functions of phasiRNAs. CONCLUSIONS: The presenting research provide insights of the molecular mechanism of phasiRNAs in regulation of GhMYB2 loci.

Gossypium barbadense and Gossypium hirsutum genomes provide insights into the origin and evolution of allotetraploid cotton.

Allotetraploid cotton is an economically important natural-fiber-producing crop worldwide. After polyploidization, Gossypium hirsutum L. evolved to produce a higher fiber yield and to better survive harsh environments than Gossypium barbadense, which produces superior-quality fibers. The global genetic and molecular bases for these interspecies divergences were unknown. Here we report high-quality de novo-assembled genomes for these two cultivated allotetraploid species with pronounced improvement in repetitive-DNA-enriched centromeric regions. Whole-genome comparative analyses revealed that species-specific alterations in gene expression, structural variations and expanded gene families were responsible for speciation and the evolutionary history of these species. These findings help to elucidate the evolution of cotton genomes and their domestication history. The information generated not only should enable breeders to improve fiber quality and resilience to ever-changing environmental conditions but also can be translated to other crops for better understanding of their domestication history and use in improvement.

LMI1-like and KNOX1 genes coordinately regulate plant leaf development in dicotyledons.

KEY MESSAGE: This report reveals that the LMI1-like and KNOX1 genes coordinately control the leaf development and different combinations of those genes which produce diverse leaf shapes including broad, lobed and compound leaves. Class I KNOTTED1-like homeobox (KNOX1) genes are involved in compound leaf development and are repressed by the ASYMMETRIC LEAVES1 (AS1)-AS2 complex. Cotton plants have a variety of leaf shapes, including broad leaves and lobed leaves. GhOKRA, a LATE MERISTEM IDENTITY 1 (LMI1)-like gene, controls the development of an okra leaf shape. We cloned the corresponding cotton homologs of Arabidopsis thaliana AS1 and AS2 and seven KNOX1 genes. Through virus-induced gene silencing technology, we found that either GhAS1 or GhAS2-silenced cotton plants showed a great change in leaf shape from okra leaves to trifoliolate dissected leaves. In the shoot tips of these plants, the expression of the cotton ortholog of Knotted in A. thaliana 1 (KNAT1), GhKNOTTED1-LIKE2/3/4 (GhKNL2/3/4), was increased. However, GhKNOX1s-silenced plants maintained the wild-type okra leaves. A novel dissected-like leaf in A. thaliana was further generated by crossing plants constitutively expressing GhOKRA with either as1-101 or as2-101 mutant plants. The dissected-like leaves showed two different leaf vein patterns. This report reveals that the LMI1-like and KNOX1 genes coordinately control leaf development, and different combinations of these genes produce diverse leaf shapes including broad leaves, lobed leaves and compound leaves. This is the first report on the artificial generation of compound leaves from simple leaves in cotton.

Identifying Functional Genes Influencing Gossypium hirsutum Fiber Quality.

Fiber quality is an important economic index and a major breeding goal in cotton, but direct phenotypic selection is often hindered due to environmental influences and linkage with yield traits. A genome-wide association study (GWAS) is a powerful tool to identify genes associated with phenotypic traits. In this study, we identified fiber quality genes in upland cotton (Gossypium hirsutum L.) using GWAS based on a high-density CottonSNP80K array and multiple environment tests. A total of 30 and 23 significant single nucleotide polymorphisms (SNPs) associated with five fiber quality traits were identified across the 408 cotton accessions in six environments and the best linear unbiased predictions, respectively. Among these SNPs, seven loci were the same, and 128 candidate genes were predicted in a 1-Mb region (±500 kb of the peak SNP). Furthermore, two major genome regions (GR1 and GR2) associated with multiple fiber qualities in multiple environments on chromosomes A07 and A13 were identified, and within them, 22 candidate genes were annotated. Of these, 11 genes were expressed [log2(1 + FPKM)>1] in the fiber development stages (5, 10, 20, and 25 dpa) using RNA-Seq. This study provides fundamental insight relevant to identification of genes associated with fiber quality and will accelerate future efforts toward improving fiber quality of upland cotton.

Genomic analyses in cotton identify signatures of selection and loci associated with fiber quality and yield traits.

Upland cotton (Gossypium hirsutum) is the most important natural fiber crop in the world. The overall genetic diversity among cultivated species of cotton and the genetic changes that occurred during their improvement are poorly understood. Here we report a comprehensive genomic assessment of modern improved upland cotton based on the genome-wide resequencing of 318 landraces and modern improved cultivars or lines. We detected more associated loci for lint yield than for fiber quality, which suggests that lint yield has stronger selection signatures than other traits. We found that two ethylene-pathway-related genes were associated with increased lint yield in improved cultivars. We evaluated the population frequency of each elite allele in historically released cultivar groups and found that 54.8% of the elite genome-wide association study (GWAS) alleles detected were transferred from three founder landraces: Deltapine 15, Stoneville 2B and Uganda Mian. Our results provide a genomic basis for improving cotton cultivars and for further evolutionary analysis of polyploid crops.

Genomic insights into divergence and dual domestication of cultivated allotetraploid cottons.

BACKGROUND: Cotton has been cultivated and used to make fabrics for at least 7000 years. Two allotetraploid species of great commercial importance, Gossypium hirsutum and Gossypium barbadense, were domesticated after polyploidization and are cultivated worldwide. Although the overall genetic diversity between these two cultivated species has been studied with limited accessions, their population structure and genetic variations remain largely unknown. RESULTS: We resequence the genomes of 147 cotton accessions, including diverse wild relatives, landraces, and modern cultivars, and construct a comprehensive variation map to provide genomic insights into the divergence and dual domestication of these two important cultivated tetraploid cotton species. Phylogenetic analysis shows two divergent groups for G. hirsutum and G. barbadense, suggesting a dual domestication processes in tetraploid cottons. In spite of the strong genetic divergence, a small number of interspecific reciprocal introgression events are found between these species and the introgression pattern is significantly biased towards the gene flow from G. hirsutum into G. barbadense. We identify selective sweeps, some of which are associated with relatively highly expressed genes for fiber development and seed germination. CONCLUSIONS: We report a comprehensive analysis of the evolution and domestication history of allotetraploid cottons based on the whole genomic variation between G. hirsutum and G. barbadense and between wild accessions and modern cultivars. These results provide genomic bases for improving cotton production and for further evolution analysis of polyploid crops.

Small interfering RNAs from bidirectional transcripts of GhMML3_A12 regulate cotton fiber development.

Natural antisense transcripts (NATs) are commonly observed in eukaryotic genomes, but only a limited number of such genes have been identified as being involved in gene regulation in plants. In this research, we investigated the function of small RNA derived from a NAT in fiber cell development. Using a map-based cloning strategy for the first time in tetraploid cotton, we cloned a naked seed mutant gene (N1 ) encoding a MYBMIXTA-like transcription factor 3 (MML3)/GhMYB25-like in chromosome A12, GhMML3_A12, that is associated with fuzz fiber development. The extremely low expression of GhMML3_A12 in N1 is associated with NAT production, driven by its 3' antisense promoter, as indicated by the promoter-driven histochemical staining assay. In addition, small RNA deep sequencing analysis suggested that the bidirectional transcriptions of GhMML3_A12 form double-stranded RNAs and generate 21-22 nt small RNAs. Therefore, in a fiber-specific manner, small RNA derived from the GhMML3_A12 locus can mediate GhMML3_A12 mRNA self-cleavage and result in the production of naked seeds followed by lint fiber inhibition in N1 plants. The present research reports the first observation of gene-mediated NATs and siRNA directly controlling fiber development in cotton.

Genetic basis for glandular trichome formation in cotton.

Trichomes originate from epidermal cells and can be classified as either glandular or non-glandular. Gossypium species are characterized by the presence of small and darkly pigmented lysigenous glands that contain large amounts of gossypol. Here, using a dominant glandless mutant, we characterize GoPGF, which encodes a basic helix-loop-helix domain-containing transcription factor, that we propose is a positive regulator of gland formation. Silencing GoPGF leads to a completely glandless phenotype. A single nucleotide insertion in GoPGF, introducing a premature stop codon is found in the duplicate recessive glandless mutant (gl2gl3). The characterization of GoPGF helps to unravel the regulatory network of glandular structure biogenesis, and has implications for understanding the production of secondary metabolites in glands. It also provides a potential molecular basis to generate glandless seed and glanded cotton to not only supply fibre and oil but also provide a source of protein for human consumption.

Sequencing of allotetraploid cotton (Gossypium hirsutum L. acc. TM-1) provides a resource for fiber improvement.

Upland cotton is a model for polyploid crop domestication and transgenic improvement. Here we sequenced the allotetraploid Gossypium hirsutum L. acc. TM-1 genome by integrating whole-genome shotgun reads, bacterial artificial chromosome (BAC)-end sequences and genotype-by-sequencing genetic maps. We assembled and annotated 32,032 A-subgenome genes and 34,402 D-subgenome genes. Structural rearrangements, gene loss, disrupted genes and sequence divergence were more common in the A subgenome than in the D subgenome, suggesting asymmetric evolution. However, no genome-wide expression dominance was found between the subgenomes. Genomic signatures of selection and domestication are associated with positively selected genes (PSGs) for fiber improvement in the A subgenome and for stress tolerance in the D subgenome. This draft genome sequence provides a resource for engineering superior cotton lines.