Cotton produces natural fiber for the textile industry. The genetic effects of genomic structural variations underlying agronomic traits remain unclear. Here, we generate two high-quality genomes of Gossypium hirsutum cv. NDM8 and Gossypium barbadense acc. Pima90, and identify large-scale structural variations in the two species and 1,081 G. hirsutum accessions. The density of structural variations is higher in the D-subgenome than in the A-subgenome, indicating that the D-subgenome undergoes stronger selection during species formation and variety development. Many structural variations in genes and/or regulatory regions potentially influencing agronomic traits were discovered. Of 446 significantly associated structural variations, those for fiber quality and Verticillium wilt resistance are located mainly in the D-subgenome and those for yield mainly in the A-subgenome. Our research provides insight into the role of structural variations in genotype-to-phenotype relationships and their potential utility in crop improvement.

Background: Verticillium wilt, caused by the fungal pathogenVerticillium dahliae, is the most severe disease in cotton (Gossypiumspp.), causing great lint losses worldwide. Disease management could be achieved in the field if genetically improved, resistant plants were used. However, the interaction betweenV. dahliaeand cotton is a complicated process, and its molecular mechanism remains obscure. To understand better the defense response to this pathogen as a means for obtaining more tolerant cultivars, we monitored the transcriptome profiles of roots from resistant plants ofG.barbadensecv. Pima90-53 that were challenged withV. dahliae.Results: In all, 46,192 high-quality expressed sequence tags (ESTs) were generated from a full-length cDNA library ofG. barbadense. They were clustered and assembled into 23126 unigenes that comprised 2661 contigs and 20465 singletons. Those unigenes were assigned Gene Ontology terms and mapped to 289 KEGG pathways. A total of 3027 unigenes were found to be homologous to known defense-related genes in other plants. They were assigned to the functional classification of plant–pathogen interactions, including disease defenses and signal transduction. The branch of "SA→NPR1→TGA→PR-1→Disease resistance" was first discovered in the interaction of cotton–V. dahliae, indicating that this wilt process includes both biotrophic and necrotrophic stages. In all, 4936 genes coding for putative transcription factors (TF) were identified in our library. The most abundant TF family was the NAC group (527), followed by G2-like (440), MYB (372), BHLH (331), bZIP (271) ERF, C3H, and WRKY. We also analyzed the expression of genes involved in pathogen-associated molecular pattern (PAMP) recognition, the activation of effector-triggered immunity, TFs, and hormone biosynthesis, as well as genes that are pathogenesis-related, or have roles in signaling/regulatory functions and cell wall modification. Their differential expression patterns were compared among mock-/inoculated- and resistant/susceptible cotton. Our results suggest that the cotton defense response has significant transcriptional complexity and that large accumulations of defense-related transcripts may contribute toV. dahliaeresistance in cotton. Therefore, these data provide a resource for cotton improvement through molecular breeding approaches.Conclusions: This study generated a substantial amount of cotton transcript sequences that are related to defense responses againstV. dahliae. These genomics resources and knowledge of important related genes contribute to our understanding of host–pathogen interactions and the defense mechanisms utilized byG. barbadense, a non-model plant system. These tools can be applied in establishing a modern breeding program that uses marker-assisted selections and oligonucleotide arrays to identify candidate genes that can be linked to valuable agronomic traits in cotton, including disease resistance.

Developing tolerant cultivars by incorporating resistant genes is regarded as a potential strategy for controlling Verticillium wilt that causes severe losses in the yield and fiber quality of cotton. Here, we identified the gene GbHyPRP1 in Gossypium barbadense, which encodes a protein containing both proline-rich repetitive and Pollen Ole e I domains. GbHyPRP1 is located in the cell wall. The transcription of this gene mainly occurs in cotton roots and stems, and is drastically down-regulated upon infection with Verticillium dahliae. Silencing HyPRP1 dramatically enhanced cotton resistance to V. dahliae. Over-expression of HyPRP1 significantly compromised the resistance of transgenic Arabidopsis plants to V. dahliae. The GbHyPRP1 promoter region contained several putative phytohormone-responsive elements, of which SA was associated with gene down-regulation. We compared the mRNA expression patterns of HyPRP1-silenced plants and the control at the global level by RNA-Seq. A total of 1735 unique genes exhibited significant differential expression. Of these, 79 DEGs involved in cell wall biogenesis and 43 DEGs associated with the production of ROS were identified. Further, we observed a dramatic thickening of interfascicular fibers and vessel walls and an increase in lignin in the HyPRP1-silenced cotton plants compared with the control after inoculation with V. dahliae. Additionally, silencing of HyPRP1 markedly enhanced ROS accumulation in the root tips of cotton inoculated with V. dahliae. Taken together, our results suggest that HyPRP1 performs a role in the negative regulation of cotton resistance to V. dahliae via the thickening of cell walls and ROS accumulation.

Verticillium wilt that is caused by Verticillium dahliae, does result in massive annual yield losses and fiber quality decline in cotton. Control by conventional mechanisms is not possible due to a wide host range and the longevity of dormant fungi in the soil in the case of absence of a suitable host. Plants have developed various mechanisms to boost their immunity against various diseases, and one is through the induction of various genes. In this research, we carried out RNA sequencing and then identified the members of the adenosine triphosphate (ATP)-binding cassette (ABC) proteins to be critical in enhancing resistance to V. dahliae infection. A total of 166 proteins that are encoded by the ABC genes were identified in Gossypium raimondii with varying physiochemical properties. A novel ABC gene, Gorai.007G244600 (ABCF5), was found to be highly upregulated, and its homolog in the tetraploid cotton Gh_D11G3432 (ABCF5), was then silenced through virus induced gene silencing (VIGS) in G. hirsutum, tetraploid upland cotton. The mutant cotton seedlings ability to tolerate V. dahliae infection was significantly reduced. Based on the evaluation of oxidant enzymes, hydrogen peroxide (H₂O₂) and malondialdehyde (MDA) showed significantly increased levels in the leaves of the mutant compared to the wild type. In addition, antioxidant enzymes, peroxidase (POD), catalase (CAT), and superoxide dismutase (SOD) concentrations were reduced in the mutant cotton leaves after treatment with V. dahliae fungi as compared to the wild type. Moreover, expression levels of the biotic stress genes, cotton polyamine oxidase (GhPAO), cotton ribosomal protein L18 (GhRPL18), and cotton polygalacturonase-inhibiting protein-1 (GhPGIP1), were all downregulated in the mutant but they were highly upregulated in the various tissues of the wild cotton seedlings. This research has shown that ABC genes could play an important role in enhancing the immunity of cotton to V. dahliae infection, and thus can be explored in developing more resilient cotton genotypes with improved resistance to V. dahliae infection in cotton.