| Plant cell walls include primary and secondary walls,which provide mechanical support and protective barriers for plant cells.The plant cell walls are mainly composed of different proportions of cellulose,hemicellulose,lignin,pectin,and a small amount of wall protein.Hemicellulose is the most abundant amorphous polysaccharide in the plant cell wall,which plays an important role in plant growth and development,stress response,and dietary nutrition.Although xylan is the principal hemicellulosic polysaccharide in grasses,the mechanism underlying its biosynthesis,in particular,the mechanism of the supply of UDP-xylose donor substrate,is largely unknown.In this study,we cloned and characterized the UXS3 gene(UDP-xylose synthase 3/UDP-glucuronic acid decarboxylase 3)in rice.The biological function of UXS3 in the xylan synthesis,cell wall remodeling,and hormone signal transduction in rice were studied.The main results are as follows:1.We identified a novel rice fragile culm mutant,named fc18(fragile culm 18).The fc18 mutant showed brittleness and severe growth defects,including stunted growth,decreased biomass,shorter panicle and grain,and decreased spike weight per plant,as compared with wild-type plants.Scanning electron microscope observation revealed that the length of cells was significantly shorter in the fc18 mutant compared to that in wild-type;2.Cell wall structure analysis revealed that the fc18 mutant showed irregular stem vascular bundles,and significantly reduced cell wall thickness.Cell wall composition determination revealed that the fc18 mutant displayed decreased levels of cellulose and hemicellulose,without significant alterations in lignin and pectin;3.The map-based cloning approach was used to isolate the fc18 locus.The fc18 locus was mapped to UXS3(UDP-glucuronate decarboxylase / UDP-xylose synthase)on chromosome 3.Sequencing of the UXS3 from fc18 identified a 24 bp nucleotide deletion in the fourth exon,resulting in a truncated FC18 protein with a substantially reduced expression level.Transgenic experiments further confirmed that the FC18/UXS3 gene is responsible for the brittleness phenotype and pleiotropic growth defects;4.Gene expression pattern and protein subcellular localization analyses showed that UXS3 gene was expressed in all tissues examined,with the highest expression level in tissues undergoing secondary wall thickening and rapid xylan accumulation.Protein subcellular location analysis revealed that the UXS3 protein is localized in the protoplasm;5.Monosaccharide analysis showed that the xylose level was decreased in fc18,and cell wall fraction determinations confirmed that the xylan content in fc18 was lower,suggesting that UDP-xylose from FC18 participates in xylan biosynthesis;6.The FC18 mutation results in cell wall remodeling,disordered orientation of cellulose microfibrils,and decreased cellulose crystallinity,which led to an enhancement in biomass saccharification;7.Transcriptome analysis showed that the differentially expressed genes of wild-type and fc18 mutants were mainly enriched in the cell wall polysaccharide metabolism,hormone signal transduction,and plant disease resistance pathways;8.We further compared the auxin content of second internodes of wild-type and fc18 plants at the heading stage by the HPLC-MS/MS assay.Compared with wild-type plants,fc18 had a lower free indole-acetic acid(IAA)content,without obvious alteration in IAA-Glu,and IAA-Asp.Gene expression analysis showed that the FC18 mutation affected the expression of PIN family genes,key genes responsible for polar auxin transport.These results suggest that the retard phenotypes of fc18 might be resulted from the decrease of IAA content.Taken together,we report the isolation of a novel rice fragile culm mutant(fc18)and the characterization of its corresponding gene,FC18/Os UXS3.Our findings demonstrate that the FC18/UXS3 plays a non-redundant role in the supply of UDP-xylose for xylan biosynthesis and is necessary for normal cellulose deposition and plant growth. |
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