GhVAS1在陆地棉棉铃脱落中的功能分析OA

[Objective]This study aimed to analyze and clarify the biological function of Gossypium hirsutum GhVAS1,and to elucidate its regulatory mechanism underlying boll abscission.[Methods]Using the Arabidopsis thaliana VAS1 gene as a query sequence,two GhVAS1 homologs(GhVAS1-A05 and GhVAS1-D05)were identified and cloned from the upland cotton cultivar TM-1.Subsequently,phylogenetic analysis,conserved motif prediction and protein structure characterization of the cloned GhVAS1 genes were performed.The spatiotemporal expression patterns of GhVAS1 were analyzed by integrating RNA-sequencing(RNA-seq)data with quantitative real-time polymerase chain reaction(qRT-PCR)analysis.The subcellular localization of GhVAS1 was determined using a transient expression system in Nicotiana benthamiana.To assess the role of GhVAS1 in boll abscission,cotton leaf crumple virus(CLCrV)-mediated virus-induced gene silencing(VIGS)was employed in cotton plants.Endogenous levels of auxin(indole-3-acetic acid,IAA)and 1-aminocyclopropane-1-carboxylic acid(ACC)in abscission layer tissues were quantified,and the transcriptional profiles of abscission-related genes were analyzed.In parallel,heterologous overexpression of GhVAS1 in A.thaliana was conducted to validate its conserved regulatory role in organ abscission across plant species.[Results]Phylogenetic analysis revealed that the two GhVAS1 share a close evolutionary relationship with AtVAS1 and encode highly conserved proteins in G.hirsutum.GhVAS1 transcripts exhibited preferential accumulation in the abscission layer and were significantly and consistently upregulated in response to abscission-inducing signals.Subcellular localization assays demonstrated that both GhVAS1 proteins are localized to the nucleus and plasma membrane.VIGS-mediated silencing of GhVAS1 resulted in a significant reduction in cotton boll abscission,which was accompanied by a marked increase in endogenous IAA content and a concurrent decrease in ACC levels in abscission layer tissues.Consistently,the transcriptional levels of key ethylene biosynthetic genes(GhACS and GhACO)and cell wall hydrolysis-related genes(GhPG and GhCEL)were significantly downregulated in GhVAS1-silenced plants.In parallel,heterologous overexpression of GhVAS1 in A.thaliana accelerated floral organ abscission compared with wild-type plants.[Conclusion]GhVAS1 positively regulates boll abscission in cotton by coordinating the IAA-ethylene metabolic balance in abscission layer tissues and upregulating the expression of cell wall degradation-related genes,thereby promoting abscission layer cell separation and subsequent organ shedding.