Molecular Mechanism Analysis Of Brassinolide Regulating The Development Of Poplar Vascular

Brassinosteroid(BR)is a sterol phytohormone that plays an important regulatory role in plant embryo development,vascular system differentiation,cell division,and plant stress resistance.The BAS1(phyB activation-tagged suppressor1)gene encodes a cytochrome P450-containing monooxygenase CYP72B1,which catalyzes the hydroxylation of the 26 th C of brassinolide to reduce the bioactivity of the hydroxylated brassinolide.It is an important brassinosteroid inactivating gene.In this study,the elevated type(BAS1-High)and dwarf type(BAS1-Dwarf)poplar trees overexpressing AtBAS1 gene were observed and the effects of endogenous brassinolide on the development of poplar vascular system were analyzed.Brassinolide(BR)and brassinolide biosynthesis inhibitor(Brz)were added to verify the effect of exogenous brassinolide on the development of poplar vascular system.Combined with gene expression level and transcriptome analysis,the mechanism of action of brassinolide on vascular development of poplar was studied,and the following results were obtained:1.Effect of brassinolide on vascular development of poplarBAS1-High compared with the control: the diameter of the myeloid cells increased by 83%,the number of cells did not differ significantly,the area of ??the pith area increased by 203%;the number of primary xylem cells increased by two layers,the number of xylem cells increased by 38%,and the diameter of xylem cells increased.50%,the area increased by 177%;the number of phloem cells increased by50%,the cell diameter did not differ significantly,the area increased by 120%;the number of xylem duct cells increased by 111%,the cell diameter increased by 77%;Compared with the control,BAS1-Dwarf reduced the diameter of myeloid cells by 45%,the number of cells by 56%,and the area of ??the medulla by 247%.The number of layers of primary xylem increased by 8 layers,the number of cells increased by 111%,and the diameter increased by 27%.The area increased by 135%;the number of phloem cells decreased by 45%,the cell diameter did not differ significantly,the phloem area decreased by 45%;the number of xylem duct cells increased by 57%,and the cell diameter increased by 25%.2.Effect of exogenous brassinolide on vascular development of poplar stemsDifferent concentrations of brassinolide can promote the growth of poplar seedlings to different extents,promote the growth of roots,and increase the lateral roots.The brassinolide synthesis inhibitor(Brz)inhibited the growth of seedlings and inhibited the growth of roots.The poplar with exogenously added brassinolide was compared with the control:the diameter of the medullary cells was increased by 80%,the number of cells was not significantly different,the area of the pith was increased by 170%;the number of layers of primary xylem increased by 3 layers,and the number of cells increased.48.9%,the cell diameter increased by 30.3%,and the area increased by 154%.The number of primary phloem cells increased by 65.5%,the cell diameter increased but not significant,the phloem area increased by 103%;the xylem duct cell diameter increased by 67%,the number increased but not significant.The poplar of the exogenous brassinolide synthesis inhibitor was compared with the control: the diameter of the medullary cells was reduced by 49%,the number was not significantly different,the pith area was reduced by 136%;the number of primary xylem cells was not significantly different,xylem The cell diameter decreased by25.7%,the number decreased slightly,the xylem area decreased by 50%;the number of primary phloem cells did not differ significantly,the cell diameter decreased by24.3%,the area decreased but there was no significant difference;the number of xylem vessels increased by 80%,cell diameter No significant difference.3.Analysis of the expression of genes related to growth and development in poplar stemsCompared with the control,the expression levels of CYP90C1 and CYP90A1 and CYP85A2 related to brassinosteroid synthesis were up-regulated in different experimental groups.The expression in Brz was lower than that in BR andBAS1-High,and the expression of CYP724B1 was Down-regulation;the expression of Brassinolide-like receptor BRI1 was up-regulated in BR and BAS1-High,and there was no significant difference in expression between Brz and BAS1-Dwarf;the regulation of cell division and elongation genes was up-regulated in BR and BAS1-High,The expression was down-regulated in Brz and BAS1-Dwarf;the xylem development-related NAC transcription factors NAC1 and NAC104 were significantly up-regulated in BR and BAS1-High,and up-regulated in Brz and BAS1-Dwarf.4.Results of transcriptome analysis of brassinosteroids on hormone metabolism and development of poplarRNA-Seq sequencing was used to compare the gene expression differences between the transgenic AtBAS1 poplar and the control: the increased and dwarfed and the control had 2701 and 738 significant differential genes(DEG),respectively.GO enrichment of differentially significant genes,classified by cell composition,molecular function,and biological processes.The biological processes are enriched by biosynthesis,cellular protein modification processes and cellular nitrogen compound metabolism.Protein components,nuclear and cell scaffolds are abundant in the cellular components.The ion function,oxidoreductase activity and kinase activity are more abundant in molecular function.The results of KEGG metabolic pathway analysis showed that the differential genes were mainly enriched in plant hormone signal transduction,carbon metabolism and starch and sucrose metabolism.The genomic hormone signal transduction pathway differential gene(Unigenes)screening results showed that there were 12 auxin-related Unigenes,9brassinosteroid-associated Unigenes,and 11 cytokinin-related Unigenes.The MYB transcription factors were initially screened: MYB3,MYB73,hormone-related genes IAA3,ARF19,CRF1 and other genes related to plant hormones and poplar vascular development.