Dissection Of Gene Regulatory Modules For Theanine Biosynthesis Based On Time Series Transcriptomic Data

As one of the three most popular non-alcoholic beverages in the world,tea contains a lot of secondary metabolites that are good for your health.Among the numerous secondary metabolites of tea trees,theanine is an important factor affecting tea quality and health as a unique non-protein amino acid.To date,only a few genes related to theanine biosynthesis has been analyzed(such as glutamine synthetase(GS),glutamine oxoglutarate aminotransferase(GOGAT),(glutamate dehydrogenase,(GDH)).Studies have shown that plant secondary metabolic pathways have modular regulatory characteristics at the molecular level,and the research on the gene regulatory modules corresponding to theanine synthesis is still blank.The analysis of these modules is of great significance for the genetic improvement of tea plants.With the development of biological sequencing technology(especially the secondgeneration and third-generation sequencing technologies),a large amount of high-quality transcriptomics data has been generated,which provides an important data foundation for the analysis of gene regulatory modules based on omics data analysis.In this paper,based on the time series transcriptomic data of theanine activation pathway,the following work is mainly carried out:(1)Theanine transcription activation treatment identified the differentially expressed genes.The theanine synthesis pathway was activated by ethylamine treatment,and transcriptome data on the activated time series were obtained.Using robust rank aggregation methods(a recognizing overlapping genes method based on rank),54 genes with consistent differential expression were identified,including 25 up-regulated genes and 29 downregulated genes article.(2)Enriched differentially expressed genes.Through functional enrichment analysis of up-regulated and down-regulated genes,it was found that the up-regulated differentially expressed genes were mainly enriched in endonuclease,hydrolase,transferase,etc.in molecular function;In biological processes,it is mainly enriched in glycyl-t RNA aminoacylation,phospholipid catabolism process,IMP metabolic process and t RNA aminoacylation;The down-regulated differentially expressed genes are mainly enriched in oxidoreductase,transferase,glutamate synthase,iron ion binding,and transmembrane transporter in molecular function;In biological processes,monoterpenoid biosynthesis process,glutathione metabolic process and ammonia assimilation cycle.(3)Gene regulatory module mining.Integrating the gene co-expression network and protein function interaction network developed by the project,thus identifying the gene function network of theanine synthesis pathway,including 45 of all(54)differentially expressed genes(83.3%)There are 127 non-redundant interactions between genes,of which there are 3 large connecting elements(overlapping each other,called gene regulation module of theanine biosynthesis)and 2 separate genes(Cs CCP interacts with Cs HASP).(4)Verification of important gene functions.Eight genes with topological importance were selected and functionally verified.The changes of theanine content were observed by high performance liquid chromatography,and the expression patterns of these 8 important topological genes were detected by Real-time Quantitative polymerase chain reaction(q RTPCR).The results show that there is a linear relationship between the theanine content change and the gene expression change,which shows that the gene regulatory module we obtained has an important relationship with theanine synthesis.In summary,theanine synthesis is a complex process that is coordinated and regulated by multiple gene regulatory modules.The results of this paper highlight the potential genes and their interaction patterns in theanine biosynthetic pathway,in order to further explore theanine.The molecular mechanism of biosynthesis and the genetic improvement of tea plants provide an important theoretical basis.