| The development and formation of xylem need a unique and complex regulation system.The important ecological value and economic value of wood make people carry out extensive research on the regulation mechanism of its development and formation.Transcriptional regulation is key regulation of xylem development and formation.Although some nodal transcription factor genes of xylem formation transcriptional regulation network have been identified and analyzed,especially the establishment of PtrSND1-Blfour-layer regulatory network of secondary cell wall biosynthesis,the overall regulatory system of secondary cell wall biosynthesis is still limited.At present,the regulatory network of secondary cell wall biosynthesis is limited.Collaterals are still in the stage of analysis and identification.Therefore,The xylem of Popoulus trichocarpa was used as the research object in this study,and tension wood(TW),which can change the composition and cell type of wood,was used as the ideal material for the study of xylem formation,13 transcription factors that may be related to the regulation of wood formation were identified by using its chromatin immunoprecipitation combined with high-throughput sequencing(ChIP-seq)and transcriptome high-throughput sequencing(RNA-seq),and compared with the vertical growth stem.Then 13 transcription factors were overexpressed in the protoplast transformation system of the stem differentiating xylem(SDX)of Popoulus trichocarpa,and 45 mainly secondary cell wall biosynthesis genes were analyzed by qRT-PCR and RNA-seq analysis to determine the regulation of 7 transcription factors PtrSND2/3-L-1,PtrMYB158,PtrMYB128,PtrMYB189,PtrMYB010,PtrMYB092 and PtrHSFB3-1on the secondary cell wall biosynthesis genes,including.11 of 19 monolignol biosynthesis genes regulated by PtrMYB092 were directly regulated,and 8 of 9 lignin monomer biosynthesis genes regulated by PtrHSFB3-1 were directly regulated.The overexpression transgenic lines and mutants of PtrHSFB3-1 reduced and increased the plant height growth,up-regulated and down regulated the expression level of 9 monolignol biosynthesis genes,increased and reduced the content of lignin,confirmed the novel regulatory function of PtrHSFB3-1,and verified the accuracy of other transcription factor regulatory conclusions.The formation of the monolignol biosynthesis enzyme protein complex can not only change the catalytic activity of the protein,but also change the catalytic properties of the protein.Understanding the formation of different protein complexes and their catalytic efficiency and characteristics is an indispensable support for the establishment of the lignin metabolism pathway model of Popoulus trichocarpa.The current model of Lignin Metabolism Pathway of Popoulus trichocarpa is based on a single enzyme catalyst,which is not accurate and complete.Only PtrC3H3,PtrC4H1and PtrC4H2,as well as Ptr4CL3 and Ptr4CL5,have been studied in 23 monolignol biosynthesis enzymes.Therefore,in this study,we tested the interaction of 23 monolignol biosynthesis enzymes one by one,and 136 of 276 combinations had interaction phenomenon.The interaction complexes of 5 membrane proteins was located on endoplasmic reticulum(ER).The interaction network of BiFC was constructed.The interaction mode of 18 monolinol biosynthesis soluble enzymes was determined by pull down mass spectrometry.The most comprehensive interaction network of monolinol biosynthesis enzymes in Popoulus trichocarpa was constructed.By analyzing the catalytic activity of PtrCCR2 and PtrCAD1,the importance of enzyme-enzyme interaction was verified.In conclusion,this study identified the regulatory role of seven transcription factors on the main biosynthesis genes of secondary cell wall,and identified the novel and unique functions of PtrHSFB3-1.The most comprehensive interaction network of 23 monolignol biosynthesis enzymes in Popoulus trichocarpa was established.All of these will provide theoretical support and guidance for molecular genetics and targeted breeding. |
PDF Full Text Request