Functional Characterization Of PtoMYB216and PtoMYB125Involved In Transcriptional Regulation Of Lignin Biosysnthesis In Populus

Lignin is the second most abundant plant biopolymer mainly present in the secondary walls of tracheary elements and fibers in wood. Lignin, cellulose and xylan constitute the secondary cell wall of plants. The lignin not only has an important role in mechanical strength, resist to pathogen attack, but also has an important economic value. In order to clarify the mechanisms of transcriptional regulation in lignin biosynthesis of P. trichocarpa, in this study, we performed sequence alignment and phylogenetic relationship between P. trichocarpa and other secondary wall-or lignin-associated MYBs in Arabidopsis thaliana, Eucalyptus gunnii and Pinus taeda, and found several lignin-associated MYBs which have a closely relationship between PtoMYBs and AtMYB58, AtMYB63, AtMYB46as well as EgMYBl. We further isolated these PtoMYBs, from P. tomentosa Carr. The expression profiles of lignin-associated PtoMYBs were investigated in various tissues of P. trichocarpa by semi-quantitative reverse-transcription (RT) PCR and quantitative real-time PCR analysis. The results reveled that PtoMYBs were expressed in various tissues and the highest levels of their transcript accumulation was observed in stems, especially for PtoMYB216and PtoMYB125. These data indicated that PtoMYBs may be involved in the biosynthesis of lignin in P. tomentosa Carr. We chose PtoMYB216and PtoMYB125for the following experiments.To determine their functions, PtoMYB216and PtoMYB125were overexpressed in transgenic Chinese white poplar (P. tomentosa Carr.) plants via Agrobacterium-mediated method. Phloroglucinol-HCl staining of the transgenic plants stem section and chemical analysis of lignin composition showed that a significant higher accumulation of lignins was detected in transgenic plants harboring the PtoMYB216and PtoMYB125overexpression construct, compared to the wild-type control, especially in transgenic PtoMYB125plants. These findings indicated that PtoMYB216and PtoMYB125may be involved in transcriptional regulation of lignin biosynthetic pathway in poplar.In order to investigate the transcriptional regulation of PtoMYBs for expression of genes involved in monolignol biosynthesis, we detected their expression levels by quantitative real-time PCR analysis. The results showed that the expression of structure genes in the lignin biosynthesis was obviously induced in transgenic plants overexpressed PtoMYB216and PtoMYB125. In addition, we also found that overexpression of PtoMYB216and PtoMYB125resulted in an increase of expression levels of these genes in cellulose synthase and xylan biosynthetic pathways.Taken together, high concentration of lignins accumulated in transgenic35S:PtoMYB216and35S:PtoMYB125plants, especially for the35S:PtoMYB125plants. Our findings indicated that PtoMYB216and PtoMYB125play an important role in transcriptional activation in the lignin biosynthesis of P. tomentosa Carr.