Transcriptional Landscape Construction Of Plant-Biotic Stress Interactions And Functional Analysis Of GhGLRs

The interaction between plants and biotic stress is a ubiquitous phenomenon in nature.Unlike plant response abiotic stresses,the interaction between plants and biotic stress factors can generate real-time information exchange and establish a complex regulatory network of molecular signals.In this study,the transcription data of the interaction between upland cotton and tobacco with pests and diseases was utilized and integrated with the transcription data of plant-biotic stress interactions from the public databases.The differential expression gene maps were constructed and used to explore transcription factors,resistance genes,and plant regulatory networks.The molecular mechanisms of the host defense system response to biotic stress were also analyzed,and an open and free database was established.At the same time,this study functionally analyzed a key gene identified in upland cotton,glutamate receptor gene（GhGLR）,that promotes defense against biotic stress and developed a batch of new disease-resistant and pest-resistant cotton materials with breeding value.The main results were as follows:1.Construction of transcription landscape of plant-biotic stress interactionsA total of 3207 transcript libraries related to biotic stresses were collected from14 plant species,including Arabidopsis,stiff brome,oilseed rape,upland cotton,soybean,barley,thistle alfalfa,tobacco,rice,broomcorn,tomato,potato,wheat and maize,from the joint public transcriptome database on plant diseases and insect pests’interactions of cotton and tobacco.The total raw data was 33.09 TB.The gene expression matrix for each plant species was constructed in response to biotic stress,which comprehensively characterized the functional gene expression profiles of plants involved in typical defense pathways and the correlation between different plant defenses.The results showed the differential expression of defense genes between dicots and monocots.91 core orthogroup were identified,providing further evidence for the broad-spectrum defense mechanisms possessed by plants.2.Identification of transcription factor regulatory networks,R genes,and co-expression networks involved in plant defense processesIn this study,we found that although the proportion of the differentially expressed gene set contained a low proportion of transcription factor families（8.19%）,it could regulate a large number of downstream genes during plant defense.We screened out 8transcription factors that respond differently to biotic stress in different plant species,and constructed the regulatory network of the transcription factor family through the expression matrix.Then,the distribution and evolution of R genes in plant species were identified,and a large number of R genes were found to be involved in the process of plant defense,as well as the APSK gene family with broad spectrum resistance was identified.Finally,the co-expression networks of different plants were constructed through the expression profiles of plants under biotic stresses,and 4 core orthogroup gene families and their related networks were identified,which also showed that they were involved in the defense system of plants through plant hormone signal transduction,MAPK signal pathway,phenylpropane biosynthesis and other pathways.3.Construction of open database of plant-biotic stress interactionsThis study visualized the differentially expressed genes involved in defense functions.As a results,we established an open database called Immunity Landscape of Plant&Biotic stress database（ILPB,http://jinlab.hzau.edu.cn/ILPB/）.The database contains 14 plant,111 different types of biotic stresses,57 plant TF families,distribution and expression of three R gene types（TNL,CNL and NL）,and information on 111,000 network core genes.The database has the following functions:1.Query of homologous genes across species.2.Functional query of genes involved in different biotic stresses of species and display pathways of gene sets.3.Identification of plant transcription factor family members and regulatory networks.4.Identification and information of R genes of species.5.Verify the co-expression network and hub genes of plant defense against different biotic stresses.4.Explore the function of the glutamate receptor gene family in upland cotton in response to biotic stressIn this study,a differentially expressed gene regulated by WRKY—glutamate receptor（GhGLR）was identified by mining and analyzing the transcriptional map of the interaction between plants and biotic stress,and 41 upland cotton species were identified by bioinformatics.The evolution and expression of GhGLRs family in biotic stresses were analyzed.It was found that the expression profiles of homologous genes in the A sub-genome and D sub-genome of the first and third subfamily were similar and found to be involved in the defense process of cotton H.armigera and S.litura.A mutant library of GhGLRs family was created using transgenic technology of upland cotton,135 independent gene-edited transgenic plants and 30 overexpression transgenic plants were obtained.Through mutant analysis,it was shown that GhGLRs were involved in the propagation of wound signals to adjacent leaves and affected the JA concentration in cotton.The GhGLRs were identified to be localized on the cell membrane and showed that the loss of GhGLR5 function reduced the Ca²⁺flow under the action of 0.1 m M glutamate solution.Finally,we used a mutant library to find that loss of GhGLR5 function reduced the resistance of upland cotton to Verticillium dahliae and S.litura.In summary,this study used large-scale transcriptional data to construct a map of differentially expressed genes across species in response to biological stress,and analyzed the responses of plant transcription factors,resistance genes,and regulatory networks to host defense responses,and further established Open database in the field of plant defense against biological stress（ILPB）.Meanwhile,the glutamate receptor protein gene family related to biotic stress in upland cotton was identified,and a mutant library was created for the family.Through mutant library analysis,the Ca²⁺signal and JA related pathways were found to be involved in the defense of upland cotton against V.dahliae and S.litura.In a word,this study promoted the research on the broad-spectrum resistance of plant defense against biotic stress,deepened the utilization of related research data,created a number of cotton materials resistant to diseases and insects with high breeding value,and greatly promoted the research on plant defense against biological stress.