The resistance to Verticillium wilt in Gossypium barbadense is generally greater than that in Gossypium hirsutum, and analyzing the differences in the mechanism and regulatory genes involved in Verticillium wilt resistance between G. barbadense and G. hirsutum is particularly important. Here, we report a transcriptomic study for phenotypic evaluation of Verticillium wilt resistance in G. hirsutum (TM-1) and G. barbadense (Hai7124) and a comparison of the transcriptomes at 7 time points after Verticillium dahliae inoculation. Phenotypic evaluation revealed that, compared with TM-1, Hai7124 was more resistant to Verticillium wilt. A total of 18138 differentially expressed genes (DEGs), including 1470 transcription factors (TFs). Further analysis of the expression of hormone biosynthesis- and signal transduction-related genes revealed that most of the genes in the salicylic acid (SA), jasmonic acid (JA) and ethylene (ET) biosynthesis pathways were highly expressed in Hai7124; the expression of SA and JA biosynthesis genes began to be significantly upregulated in the early stage of Verticillium wilt stress, while the expression of ET biosynthesis genes was upregulated mainly in the later stage. WGCNA-based binding sequence comparison revealed that MYB14 had nonsynonymous single nucleotide polymorphisms (SNPs) in 5 highly Verticillium wilt-resistant G. barbadense varieties compared to 5 highly Verticillium wilt-susceptible G. hirsutum varieties. Expression analysis revealed that GbMYB14 responded more rapidly to Verticillium wilt stress than the same gene in G. hirsutum. The resistance of G. barbadense and G. hirsutum to Verticillium wilt decreased after MYB14 silencing via virus induced gene silencing (VIGS), and leaf yellowing and necrosis in the GbMYB14-silenced plants were more obvious. Compared with those in G. hirsutum, the expression levels and of lignin biosynthesis pathway genes and the lignin content in GbMYB14-silenced plants were lower. In conclusion, our results provide a theoretical basis for an in-depth understanding of the molecular mechanism underlying the difference in Verticillium wilt resistance between G. barbadense and G. hirsutum and provide a new genetic resource for the study of cotton resistance to Verticillium wilt.

Anthocyanidin reductase (ANR) is an important regulator of flavonoid metabolism, and proanthocyanidins, the secondary metabolites of flavonoids, play an important role in the response of plants to pathogenic stress. Therefore, in this study, the expression analysis of the ANR gene family of Gossypium barbadense after inoculation with Fusarium oxysporum f. sp. vasinfectum (FOV) was performed at different time points. It was found that Gb_ANR-47 showed significant differences in the disease-resistant cultivar 06-146 and the susceptible cultivar Xinhai 14, as well as in the highest root expression. It was found that the expression of Gb_ANR-47 in the resistant cultivar was significantly higher than that in the susceptible cultivar by MeJA and SA, and different amounts of methyl jasmonate (MeJA) and salicylic acid (SA) response elements were found in the promoter region of Gb_ANR-47. After silencing GbANR-47 in 06-146 material by VIGS technology, its resistance to FOV decreased significantly. The disease severity index (DSI) was significantly increased, and the anthocyanin content was significantly decreased in silenced plants, compared to controls. Our findings suggest that GbANR-47 is a positive regulator of FOV resistance in Gossypium barbadense. The research results provide an important theoretical basis for in-depth analysis of the molecular mechanism of GbANR-47 and improving the anti-FOV of Gossypium barbadense.

Background: As a ubiquitous acid-regulating protein family in eukaryotes, 14-3-3 proteins are widely involved in the growth and development of plants. With the development of the third-generation sequencing technology and the smooth completion of the cotton genome work, it is possible to explore the existence and distribution of the 14-3-3 protein family in cotton.Results: In this paper, 33, 33, 17 and 18 members were identified from this family in Gossypium hirsutum((AD)1), G. barbadense((AD)2), G. arboreum(A2) and G. raimondii(D5), respectively. In particular, evolution analysis, structure analysis and functional expression analysis of this protein family in G. hirsutum((AD)1) were carried out. The results showed that compared with Arabidopsis and rice, the phylogenetic tree and gene structure clearly divided the 14-3-3 protein family into two subgroups in G. hirsutum((AD)1), the ε group and the non-ε group; Analysis of transcriptome expression patterns revealed that this family was significantly induced to express under abiotic stress; Most 14-3-3 proteins have a large number of cis-acting elements related to growth, development and abiotic stress in the promoter region, among which elements related to drought stress account for the largest proportion; The results of qRT-PCR showed that the expression of 14-3-3 protein had significant differences under drought stress.Conclusions: In summary, this study signified that the 14-3-3 protein family is relatively conserved in the evolutionary expansion of cotton, and may be involved in the growth and development of plants and the mechanism of stress resistance. These results provide an important theoretical and experimental basis for further analysis and verification of the function of 14-3-3 protein in cotton.

Background Domain of unknown function 668 (DUF668) may play a crucial role in the plant growth and developmental response to adverse stress. However, our knowledge of the function of the DUF668 gene family is limited. Results Our study was conducted based on the DUF668 gene family identified from cotton genome sequencing. Phylogenetic analysis showed that the DUF668 family genes can be classified into four subgroups in cotton. We identified 32 DUF668 genes, which are distributed on 17 chromosomes and most of them located in the nucleus of Gossypium hirsutum. Gene structure and motif analyses revealed that the members of the DUF668 gene family can be clustered in G. hirsutum into two broad groups, which are relatively evolutionarily conserved. Transcriptome data analysis showed that the GhDUF668 genes are differentially expressed in different tissues under various stresses (cold, heat, drought, salt, and Verticillium dahliae ), and expression is generally increased in roots and stems. Promoter and expression analyses indicated that Gh_DUF668–05, Gh_DUF668–08, Gh_DUF668–11, Gh_DUF668–23 and Gh_DUF668–28 in G. hirsutum might have evolved resistance to adverse stress. Additionally, qRT-PCR revealed that these 5 genes in four cotton lines, KK1543 (drought resistant), Xinluzao 26 (drought sensitive), Zhongzhimian 2 (disease resistant) and Simian 3 (susceptible), under drought and Verticillium wilt stress were all significantly induced. Roots had the highest expression of these 5 genes before and after the treatment. Among them, the expression levels of Gh_DUF668–08 and Gh_DUF668–23 increased sharply at 6 h and reached a maximum at 12 h under biotic and abiotic stress, which showed that they might be involved in the process of adverse stress resistance in cotton. Conclusion The significant changes in GhDUF668 expression in the roots after adverse stress indicate that GhDUF668 is likely to increase plant resistance to stress. This study provides an important theoretical basis for further research on the function of the DUF668 gene family and the molecular mechanism of adverse stress resistance in cotton.