Dissection Of The Genetic Bases Underlying The Metabolic Diversity In Rice Seeds Across Different Developmental Stages

Plant metabolites provide the material basis for plants to resist biological and abiotic stress,and also provide energy and nutrition for human.To study the metabolic diversity and the underlying genetic basis of rice is helpful to identify the regulatory genes that resist stress and improve nutrition,and provide a theoretical basis for the cultivation of stressresistant and high-nutrition varieties.Despite advance in plant metabolomics,the knowledge of the genetic and molecular basis of seed metabolome at different developmental stages is still largely unknow.To obtain a global view of the metabolic variation in rice grains of indica cultivars at different developmental stages,metabolic profiling analyses were carried out with 96 germinating seeds accessions and 108 mature seeds accessions.To visualize the accumulation pattern of these metabolites in the seeds of indica cultivars,hierarchical clustering analyses were conducted.Significant variation in the content of metabolites in germinating and mature seeds was observed within the indica accessions.The metabolic profiles of MH63 and ZS97,two representative indica rice varieties with high quality genome data,were also significantly different.To further identify the diversity in metabolite content and metabolic shifting between the seeds of MH63 and ZS97 at different developmental stages,widely targeted metabolic profiling was performed with seeds during grain filling,mature seeds and germinating seeds.855,810 and 836 metabolites were detected in seeds at three different developmental stages,among which 372 metabolites were detected in common.Principle component analysis revealed that the separation among rice grains at different stages from the same cultivar was much wider than among those of different cultivars at the same stage,indicating that the metabolic variation across developmental stages was much more significant than that between two different genotypes.To explore the genetic basis underlying the metabolic variation in seeds between MH63 and ZS97.In this study,metabolic quantitative trait loci(m QTLs)analysis was performed using recombinant inbred lines with MH63 and ZS97 as parents.A total of 4681 m QTLs were located for 2501 metabolites in seeds at three different developmental stages,and 1600,1506 and 1575 m QTLs were identified in seeds during grain filling,mature seeds and germinating seeds,respectively.When the m QTLs of individual metabolites were compared,we detected 1,804 distinct loci among the total 2068 loci for the 372 codetected metabolites in the seeds at three different stages,suggesting that the majority of metabolites across the three stages under different genetic control.Further analysis of the m QTLs of the three developmental stages,we found that the genetic regulation of metabolite accumulation was closely related to the developmental stage.To identify candidate genes underlying the metabolic variation in the seeds of MH63 and ZS97,in silico analyses with genomic,transcriptomic and metabolomic data were carried out.Data mining identified 36 candidate genes modulating levels of metabolites,among which LOC?Os06g03990 may affect the accumulation of asparagine.We confirm the in vivo function of LOC?Os06g03990 by transgenic experiment.Through analyzing the correlation between metabolites in different developmental seeds,we found that compounds of the same chemical class or those involved in the same biochemical pathway tended to display tight correlations with each other.Metaboliteagronomic trait association and colocation between m QTLs and phenotypic quantitative trait loci(p QTLs)revealed the complexity of the metabolite-agronomic trait relationship and the corresponding genetic basis.In order to compare the difference of beneficial metabolites between MH63 and ZS97,metabolic profiling of polished rice and brown rice was performed.Further anlysis of the metabolome of brown rice and polished rice found that the content of metabolites in brown rice were more than 1.5 times higher than those in polished rice,including 80.0% amino acids,56.7% vitamins and 82.3% flavonoids,indicating that brown rice is more nutritious.This study significantly improved our knowledge of the genetic and biochemical basis of rice seed metabolome variation in different stages and provides profound insights into crop improvements.