Functional Analysis Of PxNAC73 Gene During Wood Formation In Populus × Xiaohei T. S. Hwang Et Liang

NAC transcription factor is a kind of plant-specific transcription factor,which participates in many biological processes,such as the development of floral organs and the establishment of embryonic organs.Previous studies had shown that NAC transcription factor also played an important role in the development and formation of secondary cell wall.In this study,a NAC transcription factor Px NAC73 was screened out from the stem transcriptomics data of Populus× xiaohei treated with nitrogen.The gene was a nitrogen-responsive gene,and its expression was significantly increased under low nitrogen treatment.Meanwhile,its expression was also significantly changed at different stages of stem development,predicting that the gene might play a role during stem development.Therefore,P.× xiaohei was used as the plant material to study the functions of Px NAC73 transcription factor in the wood formation process.The main results were as follows:(1)The transcription factor Px NAC73 was highly expressed in secondary stem by RTq PCR.The full-length sequence of Px NAC73 CDS was cloned from the c DNA of P.× xiaohei.Subcellular localization indicated that Px NAC73 protein was localized in nucleus,and autoactivation activity of transcription factors analysis indicated that the protein had transcriptional activation activity.(2)The p ROKⅡ-Px NAC73 recombinant expression vector and the p ROKⅡ-Px NAC73-SRDX dominant inhibition expression vector were constructed,and genetically transformed into P.× xiaohei.Px NAC73-SRDX dominant inhibition expression lines were obtained.(3)The growth phenotype analysis showed Px NAC73-SRDX lines grew more slowly and shorter than WT.The observation under the scanning electron microscope showed that the fiber cell wall of transgenic lines was significantly thinned.Histochemical staining analysis showed that the secondary xylem width of transgenic lines was decreased and xylem development was inhibited.Besides,the length and diameter of fibroblasts in the transgenic lines were significantly reduced.RT-q PCR results of secondary wall synthesis related genes showed that the expression levels of most genes(16)in the lignin synthesis pathway were significantly reduced,and the expression levels of two genes in the cellulose synthesis pathway were significantly reduced.The results of wood component analysis showed that the lignin and cellulose contents of transgenic lines were significantly reduced,and the differences between hemicellulose content was not significantly.However,the content of sugar content was determined and analyzed,and it was found the contents of reducing sugar and starch in the transgenic lines were increased significantly.The RT-q PCR results showed that the expression levels of genes related to sucrose metabolic pathways were not significantly changed,while the expression levels of genes related to starch metabolic pathways were significantly increased.In summary,PxNAC73 was a transcription factor belonging to NAC family with transcriptional activation activity,which was located in the nucleus.It inhibited the biosynthesis of lignin and cellulose,thereby affecting the formation of secondary wall and resulting in the slow development of secondary xylem.These results could lay experimental materials for further study on the function of Px NAC73 gene in the formation of secondary xylem of P.× xiaohei,and also provide theoretical foundation for studying the mechanism of wood formation in forest trees.