Profiles Of MicroRNA Expressed During Regeneration Of The Secondary Vasular System In Poplar(Populus Tomentosa Carr.)

Wood formation is a complex progress composing many biological events,occurs as a result of growth and differentiation of the vascular cambium. This secondary growth is a highly ordered developmental progress,which coordinates functional genes and different regulators. MicroRNAs (miRNAs), a group of small non-coding RNAs, are a class of post-transcriptional negative regulators playing vital roles in plant development and growth. To investigate miRNAs involved in secondary vascular system (SVS) development, aμParaflo? MicroRNA microarray was used to analyze miRNAs expression profiles during regeneration of SVS in Chinese white poplar (Populus tomentosa).By comparing the expressional profiles of adjacent regeneration stages, 99 differentially expressed miRNAs were identified, most of them belong to the superfamilies of miRNA159, miRNA164, miRNA166, miRNA319, miRNA396, miRNA168 and miRNA399. Cluster analysis showed that the expression pattern of 76 miRNAs changed dramatically during SVS formation. Those miRNAs also belong to the seven miRNAs superfamilies mentioned above. Previous data proved that the targets of these miRNAs are transcriptional factors play important roles in wood formation. Thus, we can suggest that some members of these miRNA families might be the negative regulators of the SVS in poplar. And those expressed differentially on 12th, 14th, 16th and 22nd day of the regeneration might involve in the stage switches during SVS regeneration. The expression pattern of several miRNAs was confirmed by real time quantitive PCR.To characterize the function of miRNA target genes, 7 Arabidopsis T-DNA mutant lines of MYB were chosen to further analysis according to the previous data on their differentially expression during SVS in poplar. The mutants and wild type(WT) Arabidopsis were cultured under secondary growth inducing condition. An Arabidopsis mutant named CWT had obviously changes in interfascicular regions, where the secondary walls of homozygous cwt mutant were clearly thickened, the cell cavity and quantity of interfascicular fiber reduced relatively. Secondary walls of xylem thickened equally but gradually in WT lines.miRNAs profiling during SVS regeneration were performed by liquid miRNAs chip in this study. The results showed that several miRNAs might play important roles in cambium development and xylem differentiation. And mutant analysis of MYB, the target gene of miRNA159, showed the irregular cell wall thickness in certain region, which suggested this miRNA might regulate the formation of secondary wall. Our data might provide new insight into the transcriptional regulatory networks of wood formation, and select candidate genes may be responsible for the traits of wood property. To isolate further and characterize these key genes is needed to understand their biological function in wood formation in a molecular breeding point of view.