The OsmiRNA166b-OsHox32 pair regulates mechanical strength of rice plants by modulating cell wall biosynthesis

The plant cell wall provides mechanical strength to support plant growth and development and to determine plant architecture. Cellulose and mixed-linkage glucan (MLG) present in primary cell wall, whereas cellulose, lignin and hemicellulose exist in secondary cell wall. Biosynthesis of the cell wall biopolymers needs the coordinated transcriptional regulation of all the biosynthetic genes. The module of OsmiR166b-OsHox32 regulates expression levels of the genes related to biosynthesis of MLG, cellulose and lignin. Transgenic plants knocking down miR166b (STTM166b) by short tandem target mimic (STTM) technology or overexpressing OsHox32 (OEHox32) showed drooping leaves and brittle culms. Due to accumulation of less lignin and cellulose, the cell wall thickness of STTM166b and OEHox32 plants was reduced when compared to that of wild-type plants. Overexpression of miR166b (OE166b) in rice plants or knocking down of OsHox32 by RNA interference (RNAiHox32) led to increased thickness of cell walls and enhanced mechanical strength of culms. Molecular analyses showed that OsmiR166b-OsHox32 pair regulates cell wall-related gene expression. OsHox32 binds to the promoters of OsCAD2 and OsCESA7 to suppress the expression levels of these two genes. The suppression of OsCAD2 is synergistic when OsHox32 is co-expressed with OSH15 (Oryza sativa homeobox 15). OsHox32 interacts with OSH15, and the START domain of OsHox32, harbouring the miR166b cleavage site, is required for the interaction of these two proteins. Our results demonstrate that OsmiR166b-OsHox32 pair plays important roles not only in plant growth and development but also in plant architecture by regulating the cell wall-related gene expression.