Functional Characterization Of Poplar PtrMYB-like

The MYB family of proteins is large, functionally diverse and represented in all eukaryotes. In plants, the MYB family has selectively expanded, particularly through the large family of R2R3-MYB. Numerous R2R3-MYB proteins have been found to be involved in the control of plant-specific processes including primary and secondary metabolism, cell fate and identity, developmental processes and responses to biotic and abiotic stresses. Plant phenylpropanoid-derived compounds are a large family of phenylalanine-derived secondary metabolites that includes monolignols, flavonoids, phenolic acids, and these compounds have roles in plant growth and development. Many reports focus on the regulation of R2R3-MYB in one secondary metabolites biosynthesis (such as lignin or anthocyanin). However, little is known on the function of MYB-related protein only containing single MYB domain in plants. Here we report a PtrMYB-like gene from Populus trichocarpa, which plays a double role in anthocyanin accumulation and secondary cell wall formation. The main results are as follows:The Locus Name of PtrMYB-like is Potri.015G069000 containing 3 exons and 2 introns. The full-length sequence of CDS consists of 954 nucleotides encoding 318 amino acids. Homology blast and sequence analysis shows that poplar PtrMYB-like homologues exist in plants such Arabidopsis thaliana, Triticum aestivum, Glycine max, and these genes are highly conservative in phylogeny.Quantitative RT-PCR results indicate that PtrMYB-like is expressed in all the examined tissues, and most strongly in xylem, cambium, phloem, young leaf, mature leaf and petiole but weaker in apical bud and root. Moreover, the transcriptional level of PtrMYB-like continuously increases from IN1 to IN4 stem, and are different degree in IN5 to IN 10 stem.We prepare a C-terminal fusion of GFP to PtrMYB-like and place it under the control of the CaMV 35S promoter, then the construct is transformed into Arabidopsis. We monitor GFP fluorescence in the roots of transgenic Arabidopsis and find that PtrMYB-like is localized to the nucleus.PtrMYB-like overexpression leads to anthocyanin accumulation in 4-day-old seedings of transgenic Arabidopsis. The expression levels of structural genes and regulatory genes involved in anthocyanin biosynthesis increase at different degrees, suggesting PtrMYB-like regulates anthocyanin biosynthetic pathway. Furthermore, PtrMYB-like overexpression plants cause a slow growth phenotype in 25-day-old seedings. PtrMYB-like overexpression plants xylary fibers appear to have a collapsed morphology, and the expression levels of SCW-related genes are elevated to different degree, indicating PtrMYB-like is involved in secondary cell wall formation in Arabidopsis.Meanwhile, the reverse genetics technology is used to specifically downregulate PtrMYB- like gene expression in Populus 84K(Populus alba x P. glandulosa). We produce highly efficient gene-silencing transgenic poplars, laying the foundation for analyzing the function of PtrMYB-like in Populus.