The Mechanism Of Interaction Between Opaque2 And ZmGRAS11 To Regulate Maize Starch Synthesis

Maize(Zea mays L.)is the frist major crops in China,and it's also a vital food,feed and industrial raw material.Starch is an important part of maize kernels.The total amount of starch and the ratio of amylose to amylopectin affect the yield,edible quality and industrial quality of maize.The synthesis of starch is coordinated by a variety of enzymes,including sucrose synthase(SUS),ADPG pyrophosphorylase(AGPase),starch synthase(SSs),starch branching enzyme(SBE),and starch debranching enzyme(DBE).Opaque2(O2)protein plays an important role in maize endosperm growth and metabolism,the grain starch content and yield decreased significantly in o2 mutant compared to the control.Previous studies have shown that O2 protein directly regulates the key enzymes of starch synthesis SUS and SSs,and indirectly regulates SBE to participate in starch synthesis.Many genes are related to starch synthesis that regulated by multiple transcription factors at the transcription level.Previous analysis of the RNA-Seq data revealed that the expression of the transcription factor ZmGRAS11 was dramatically decreased in the o2 mutants.It is speculated that ZmGRAS11 may participate in starch accumulation together with O2 protein by regulating starch synthesis-related genes directly.Through cluster analysis,qRT-PCR,subcellular localization,yeast one-hybrid and yeast two-hybrid,we use D Huang 212 and D Huang 212/o2 to analyze the regulatory role of ZmGRAS11 in the starch synthesis pathway,and explore the interaction mechanism between O2 and ZmGRAS11 initially.The main results are as follows: 1.Phylogenetic tree analysis showed that ZmGRAS11 belongs to the DELLA subfamily of the GRAS family,and its homologous genes are involved in the endosperm development in rice.Compared with the control,the expression of ZmGRAS11 in the endosperm of the o2 mutant was significantly reduced from 10 to 26 days after pollination.2.GRAS belongs to a transcription factor's family.ZmGRAS11 is found to be a nuclear protein with subcellular localization,and it has self-activating activity.Using the yeast one-hybrid system,we found that ZmGRAS11 can bind to the promoter regions of the key starch synthesis genes Sh1,SSI,SSIIIa,and SBEIIb.3.Yeast two-hybrid assay showed that ZmGRAS11 interacts with O2 protein.We speculate that ZmGRAS11 participate in starch synthesis together with O2 protein.This study is facilitated to further improve the research on the regulation mechanism of starch synthesis in o2 mutants,and to provide theoretical guidance for molecular breeding of high-quality protein maize.