Modulating Starch Metabolism By A Novel Rice Transcription Factor OsSGL

Rice is one of the three major food crops on earth and is an important food resource in China.With the improvement of living standards,people are no longer satisfied with having enough food and clothing,but desire to have high quality food.In order to meet this goal,efforts have been made on to improve the quality of rice,et al.The main edible part of rice grain is the endosperm,of which starch is the main component.The content and structure of starch are of the most important factors in determining the steaming and palatability quality of rice grain.Although the starch synthesis-related genes(SSRG)are well studied,the regulation of the expression of SSRG-related genes has received little attention.In this study,a novel gene involving in rice stress tolerance and grain length(OsSGL)was studied.The OsSGL over-expression(OE)and RNA interference(RNAi)mutant plants were generated and the grains harvested from these plants were analyzed to study the effects and possible mechaaaanisms of OsSGL on grain quality.The main findings were as follows.(1)For the first time,OsSGL was found to be involved in the regulation of starch metabolic pathways.Compared with wild-type SR59,the starch granules in the abdomen of the endosperm of OsSGL-OE grains were small and loosely arranged.The relative crystallinity and thermodynamic properties of starch were not altered in the OsSGL-OE and interference grains,but their peak and final viscosities were lower than those of the wild type.(2)OsSGL protein localized in the nucleus and had transcriptional activity.ChIP-Seq analysis revealed that OsSGL was able to bind 6900 chromatin regions in OsSGL-OE plants,particularly enriched in the TSS region.KEGG analysis of the genes in the bound regions revealed that the enriched genes were involved in the sucrose and starch metabolic signalling pathway.The binding of OsSGL protein to the promoter of the sucrose synthase 1 gene(OsSUS1)was demonstrated by yeast monohybrid,ChIP,dual-luciferase LUC and EMSA assay.This binding suppressed OsSUS1 transcription,demonstrating that OsSGL is a novel transcription factor that regulates starch metabolism.(3)To further investigate the roles of OsSGL in regulating starch metabolism,yeast sieve library was screened to identify OsSGL binding protiens.The result showed that OsSGL interacted directly with OsGAPC3,a key rate-limiting enzyme of glycolysis.This interaction was verified by yeast two-hybrid,BiFC,GST-pull down and Co-IP experiments.OsGAPC3-OE rice was generated and the grains were analyzed.The results showed that,when compared with wild-type Japanese Haru,the chalkiness of OsGAPC3-OE grain increased,the total starch content decreased and the total protein content increased,but grain length and width did not change,suggesting that OsSGL may regulate the starch metabolism process through the glycolytic pathway.