Regulation Of PtTDIFs Gene Family On Vascular Tissue Development

The tracheary element differentiation inhibitory factor(TDIF)plays a crucial role in the regulation of vascular system development.The endogenous TDIF peptide of Arabidopsis thaliana is encoded by CLAVATA3/EMBRYO SURROUNDING REGION 41(CLE41)and CLE44 genes,while the TDIF and TDIF-like peptides in poplar are encoded by the CLE genes PtTDIF1,2,3,4 and PtTDIF-like1,2 respectively.There is only one amino acid variation in the 12 amino acid CLE conserved domain of TDIF and TDIF-like peptides.To investigate the dependency of the in vivo function of PtTDIFs on the 12 amino acid CLE conserved domain,A.thaliana seedlings were treated with in vitro chemically synthesized TDIF and TDIF-like peptides,and it was found that the two peptides had similar effects on multiple aspects of vascular system development in herbaceous model plants.However,morphological and histological section analysis showed that overexpression of four genes had significant differences in 35S:PtTDIF2,35S:PtTDIF3,35S:PtTDIF-like1,and 35S:PtTDIF-like2 transgenic lines.Compared with PtTDIF3 and PtTDIF-like2 genes,overexpression of PtTDIF2 and PtTDIF-like1 genes had much more pronounced effects on the morphological structure of Arabidopsis vascular tissue,suggesting that the in vivo function of PtTDIFs genes is not completely dependent on 12 peptide CLE conserved domains.In addition,the expression levels of WUSCHEL-related HOMEOBOX 4(WOX4)and WOX14 significantly increased in 35S:PtTDIFs transgenic Arabidopsis hypocotyl and decreased in 35S:PtTDIFs transgenic Arabidopsis stem,revealing that the difference of vascular cambium cell division activity may be related to the differential expression of the WOX gene in the two tissues.To further explore the factors influencing the in vivo function of PtTDIFs,we continued to use Agrobacterium tumefaciens containing PtTDIFspro:GUS or 35S:PtTDIFs expression vectors to transform INRA717 1-B4 poplars.The genome DNA and semi-quantitative PCR data confirmed that the relevant transgenic plants were obtained and morphological analyses showed that the height and stem diameter of 35S:PtTDIFs transgenic poplars were both reduced.In summary,this study discussed the function of the PtTDIFs gene family in the regulation of plant vascular tissue development and laid a foundation for in-depth study of the biological functions of the CLE gene family in plants.