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Branch length is an important plant architecture trait in cotton (Gossypium) breeding. Development of cultivars with short branch has been proposed as a main object to enhance cotton yield potential, because they are suitable for high planting density. Here, we report the molecular cloning and characterization of a semi-dominant quantitative trait locus, Short Branch Internode 1(GhSBI1), which encodes a NAC transcription factor homologous to CUP-SHAPED COTYLEDON 2 (CUC2) and is regulated by microRNA ghr-miR164. We demonstrate that a point mutation found in sbi1 mutants perturbs ghr-miR164-directed regulation of GhSBI1, resulting in an increased expression level of GhSBI1. The sbi1 mutant was sensitive to exogenous gibberellic acid (GA) treatments. Overexpression of GhSBI1 inhibited branch internode elongation and led to the decreased levels of bioactive GAs. In addition, gene knockout analysis showed that GhSBI1 is required for the maintenance of the boundaries of multiple tissues in cotton. Transcriptome analysis revealed that overexpression of GhSBI1 affects the expression of plant hormone signalling-, axillary meristems initiation-, and abiotic stress response-related genes. GhSBI1 interacted with GAIs, the DELLA repressors of GA signalling. GhSBI1 represses expression of GA signalling- and cell elongation-related genes by directly targeting their promoters. Our work thus provides new insights into the molecular mechanisms for branch length and paves the way for the development of elite cultivars with suitable plant architecture in cotton., (© 2024 The Author(s). Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Background: BRVIS RADIX (BRX) family is a small gene family with the highly conserved plant-specific BRX domains, which plays important roles in plant development and response to abiotic stress. Although BRX protein has been studied in other plants, the biological function of cotton BRX-like (BRXL) gene family is still elusive., Result: In this study, a total of 36 BRXL genes were identified in four cotton species. Whole genome or segmental duplications played the main role in the expansion of GhBRXL gene family during evolutionary process in cotton. These BRXL genes were clustered into 2 groups, α and β, in which structural and functional conservation within same groups but divergence among different groups were found. Promoter analysis indicated that cis-elements were associated with the phytohormone regulatory networks and the response to abiotic stress. Transcriptomic analysis indicated that GhBRXL2A/2D and GhBRXL5A/5D were up/down-regulated in response to the different stress. Silencing of GhBRXL5A gene via virus-induced gene silencing (VIGS) improved salt tolerance in cotton plants. Furthermore, yeast two hybrid analysis suggested homotypic and heterotypic interactions between GhBRXL1A and GhBRXL5D., Conclusions: Overall, these results provide useful and valuable information for understanding the evolution of cotton GhBRXL genes and their functions in salt stress., (© 2024. The Author(s).)

C-TERMINALLY ENCODED PEPTIDEs ( CEPs ) are a class of peptide hormones that have been shown in previous studies to play an important role in regulating the development and response to abiotic stress in model plants. However, their role in cotton is not well understood. In this study, we identified 54, 59, 34, and 35 CEP genes from Gossypium hirsutum (2n = 4x = 52, AD1), G. barbadense (AD2), G. arboreum (2n = 2X = 26, A2), and G. raimondii (2n = 2X = 26, D5), respectively. Sequence alignment and phylogenetic analyses indicate that cotton CEP proteins can be categorized into two subgroups based on the differentiation of their CEP domain. Chromosomal distribution and collinearity analyses show that most of the cotton CEP genes are situated in gene clusters, suggesting that segmental duplication may be a critical factor in CEP gene expansion. Expression pattern analyses showed that cotton CEP genes are widely expressed throughout the plant, with some genes exhibiting specific expression patterns. Ectopic expression of GhCEP46-D05 in Arabidopsis led to a significant reduction in both root length and seed size, resulting in a dwarf phenotype. Similarly, overexpression of GhCEP46-D05 in cotton resulted in reduced internode length and plant height. These findings provide a foundation for further investigation into the function of cotton CEP genes and their potential role in cotton breeding.

Background: Members of the AT-HOOK MOTIF CONTAINING NUCLEAR LOCALIZED (AHL) family are involved in various plant biological processes via protein-DNA and protein-protein interaction. However, no the systematic identification and analysis of AHL gene family have been reported in cotton., Results: To investigate the potential functions of AHLs in cotton, genome-wide identification, expressions and structure analysis of the AHL gene family were performed in this study. 48, 51 and 99 AHL genes were identified from the G.raimondii, G.arboreum and G.hirsutum genome, respectively. Phylogenetic analysis revealed that the AHLs in cotton evolved into 2 clades, Clade-A with 4-5 introns and Clade-B with intronless (excluding AHL20-2). Based on the composition of the AT-hook motif(s) and PPC/DUF 296 domain, AHL proteins were classified into three types (Type-I/-II/-III), with Type-I AHLs forming Clade-B, and the other two types together diversifying in Clade-A. The detection of synteny and collinearity showed that the AHLs expanded with the specific WGD in cotton, and the sequence structure of AHL20-2 showed the tendency of increasing intron in three different Gossypium spp. The ratios of non-synonymous (Ka) and synonymous (Ks) substitution rates of orthologous gene pairs revealed that the AHL genes of G.hirsutum had undergone through various selection pressures, purifying selection mainly in A-subgenome and positive selection mainly in D-subgenome. Examination of their expression patterns showed most of AHLs of Clade-B expressed predominantly in stem, while those of Clade-A in ovules, suggesting that the AHLs within each clade shared similar expression patterns with each other. qRT-PCR analysis further confirmed that some GhAHLs higher expression in stems and ovules., Conclusion: In this study, 48, 51 and 99 AHL genes were identified from three cotton genomes respectively. AHLs in cotton were classified into two clades by phylogenetic relationship and three types based on the composition of motif and domain. The AHLs expanded with segmental duplication, not tandem duplication. The expression profiles of GhAHLs revealed abundant differences in expression levels in various tissues and at different stages of ovules development. Our study provided significant insights into the potential functions of AHLs in regulating the growth and development in cotton.

Introduction: The phosphatidylethanolamine-binding protein (PEBP) gene family plays a crucial role in seed germination, reproductive transformation, and other important developmental processes in plants, but its distribution in Gossypium genomes or species, evolutionary properties, and the fates of multiple duplicated genes remain unclear., Objectives: The primary objectives of this study were to elucidate the distribution and characteristics of PEBP genes in Gossypium, as well as the evolutionary pattern of duplication and deletion, and functional differentiation of PEBPs in plants., Methods: Using the PEBP protein sequences in Arabidopsis thaliana as queries, blast alignment was carried out for the identification of PEBP genes in four sequenced cotton species. Using the primers designed according to the PEBP genome sequences, PEBP genes were cloned from 15 representative genomes of Gossypium genus, and the gene structure, CDS sequence, protein sequence and properties were predicted and phylogenetic analysis was performed. Taking PEBP proteins of grape as reference, grouping of orthologous gene, analysis of phylogeny and divergence of PEBPs in nine species were conducted to reconstruct the evolutionary pattern of PEBP genes in plants., Results: We identified and cloned 160 PEBPs from 15 cotton species, and the phylogenetic analysis showed that the genes could be classified into the following three subfamilies: MFT-like, FT-like and TFL1-like. There were eight single orthologous group (OG) members in each diploid and 16 double OG members in each tetraploid. An analysis of the expression and selective pressure indicated that expression divergence and strong purification selection within the same OG presented in the PEBP gene family., Conclusion: An evolutionary pattern of duplication and deletion of the PEBP family in the evolutionary history of Gossypium was suggested, and three pairs of genes resulted from different whole-genome duplication events.

Key Message: Nulliplex-branch (nb) mutants in cotton display a specific architecture. The gene responsible for the nb phenotype was identified, and its modulation mode was further studied. Plant architecture is an important agronomic factor influencing various traits such as yield and variety adaptability in crop plants. Cotton (Gossypium) simultaneously displays monopodial and sympodial growth. Nulliplex-branch (nb) mutants showing determinate sympodial shoots have been reported in both G. hirsutum (Ghnb) and G. barbadense (Gbnb). In this study, the gene responsible for the nb phenotype was identified. GhNB and GbNB were found to be allelic loci and are TERMINAL FLOWER 1 orthologs on the Dt subgenome, though the At copies remain native. Sequencing and association analyses identified four (Gh-nb1-Gh-nb4) and one (Gb-nb1) type of point mutation in the coding sequences of Ghnb and Gbnb, respectively. The NB gene was mainly expressed in the root and shoot apex, and expression rhythms were also observed in these tissues, suggesting that the expression of the NB gene could be regulated by photoperiod. Constitutive overexpression of GhNB suppresses the differentiation of the reproductive shoots. Knockout of both copies of GhNB caused the main and lateral shoots to terminate in flowers, which is a more determinate architecture than that of the nb mutants and implies that its function might be dosage dependent. A protein lipid overlay assay indicated that the amino acid substitutions in Gh-nb1 and Gb-nb1 weaken the ligand-binding activity of the NB protein in vitro. These findings suggest that the NB gene plays crucial roles in regulating the determinacy of shoots, and the modulation of this gene should constitute an effective crop improvement approach through adjusting the growth habit of cotton.