榧树小孢子叶球快速伸长和开裂的分子调控

[Objective]Exploring the molecular mechanisms of rapid elongation and final dehiscence during the development of staminate strobilus in Torreya grandis.[Method]From February to April,we collected samples every 7 days for a total of 8 periods.Staminate strobilus samples at eight stages underwent morphological measurement and transcriptome sequencing.Key modules and genes are extracted by WGCNA analysis by fusing transcriptome data with morphological traits.Then,to confirm their function,the main genes'transient expression in the T.torreya's staminate strobilus was utilised.Finally,the regulatory function of important genes was confirmed using WGCNA correlation analysis in conjunction with luciferase assays and yeast one hybrid.[Result](1)T.grandis'staminate strobiles were measured for their shape,and it was discovered that they went through three stages of growth:gradual growth from XBZ1 to XBZ5,rapid elongation from XBZ5 to XBZ8,and dehiscence during XBZ8.(2)Six modules relevant to rapid elongation were identified by WGCNA analysis of the transcriptome and morphological data;the turquoise module had the highest number of connected genes,including transcription factors like WOX and PIF4,and it was used to create a transcription factor regulatory network.(3)When compared to the control,transient expression of TgPIF4 in staminate strobilus dramatically improved staminate strobilus elongation.Potential TgPIF4 target genes were discovered using WGCNA association analysis.TgPIF4 was discovered to bind to the promoters of TgTIR2 and TgNADP-ME to regulate their expression and promoted the rapid elongation of staminate strobiles in T.grandis,according to the results of luciferase assays,yeast one hybrid and RT-PCR.(4)Jasmonic acid,methyl jasmonate,lignin,and lignin monomer levels increased considerably in the staminate strobilus during the dehiscence stage.They were most closely associated to the red module according to WGCNA analysis.At the dehiscence stage,17 and 15 genes from the phenylpropanoid pathway and linolenic acid metabolism,respectively,increased considerably in the red module.(5)Through the dual luciferase assay,TgWRKY1 and TgWRKY2 were screened to activate the LUC activity driven by TgPAL and TgJMT promoters,respectively.Yeast one hybrid experiment findings also demonstrated that TgWRKY1 and TgWRKY2 could bind to P1 promoter of TgPAL and P2 promoter of TgJMT,respectively.Transient expression of TgWRKY1 and TgWRKY2 genes in staminate strobiles can respectively enhance the expression of TgPAL and TgJMT genes,resulting in an increase in the concentration of lignin and Methyl jasmonate.This increase in concentration can promote the dehiscence of staminate strobiles.[Conclusion]This study discovered that TgPIF4 can regulate the expression of TgTIR2 and TgNADP-ME during the rapid elongation stage,promoting the rapid elongation of T.grandis'staminate strobilus.TgWRKY1 and TgWRKY2 can regulate the expression of TgPAL and TgJMT during the dehiscence stage,increase the concentration of lignin and methyl jasmonate,and promote the dehiscence of staminate strobilus in T.grandis.