Functional Study Of Different Alternative Splicing Of Plant Glycosyltransferase SEC Gene

In this research,we studied the biological functions of different splices of glycosyltransferase SEC in Arabidopsis thaliana O-GlcNAc modification and the heterologous gene SEC in Fagopyrum esculentum.In Arabidopsis thaliana,the long splice At SEC.1 and the short splice At SEC.2 with 6 exons difference were produced by the alternative splicing event of At SEC gene.The Fe SEC1 and Fe SEC2 glycotransferase genes of Fagopyrum esculentum with highly similar sequence At SEC.1 and At SEC.2 were obtained by the transcriptomes data of Fagopyrum esculentum in our lab,respectively.The transcriptional expression patterns of At SEC.1,At SEC.2,Fe SEC1 and Fe SEC2 were analyzed by fluorescence quantitative q PCR assay.It was found that the expression of At SEC.1 was the highest in mature flowers,and the abundance of At SEC.2 was the highest in silique.In Fagopyrum esculentum,Fe SEC1 and Fe SEC2 were all highly expressed in pistil.Through cloning,multiple sequence alignment and domain analysis,it is shown that At SEC.1,At SEC.2,Fe SEC1 and Fe SEC2 all have TPRs domains.The evolutionary tree was further constructed to illustrate the evolutionary relationship between these four genes and their homologous genes.In this subject,the function of At SEC.1,At SEC.2,Fe SEC1 and Fe SEC2 in Arabidopsis thaliana was studied.Compared with the wild type,the functional expression of At SEC.1 in Arabidopsis thaliana led to the phenomenon of incomplete petal development,stamen disappear,gynoecium malformation with exposed ovule and shorter fruit length.35S::At SEC.2-GFP Arabidopsis thaliana transgenic lines only the stamens could not fully develop,and the four strong stamens become weak.In 35S::Fe SEC1-GFP transgenic Arabidopsis thaliana lines,the four sepals were two long and two short,the number of pistils was increased with ectopic expression,the stigmas and ovule structures were obvious,and some plants showed six petals.No obvious phenotype was observed in 35S::Fe SEC1-GFP transgenic lines.The silencing of At SEC.1 and At SEC.2 splices based on TRV-VIGS technique can lead to earlier flowering time and disorderly flower development in Arabidopsis thaliana.Compared to the control group,the development of petals and pistils of Arabidopsis thaliana treated with p TRV2-At SEC.1 and p TRV2-At SEC.2 was inhibited,and the whole flower was aborted.Compared with the silencing of At SEC.2 gene,the silencing of At SEC.1 gene had no effect on pistil.Based on the analysis of multiple transcriptome data,it was found that the Arabidopsis glycosyltransferase SEC was induced by Pseudomonas syringae,which may be related to the basic resistance of Arabidopsis.It was proved that the At SEC.1 and At SEC.2 splice in Arabidopsis thaliana were most induced 24 h after Pseudomonas syringae infection.Apply SA and Pip to wild-type Arabidopsis to verify the role of Arabidopsis SEC in SA and Pip signal pathways.It was found that At SEC.1 may act in the downstream of SA and Pip synthesis pathways,while At SEC.2 acts on the downstream of SA,not induced by Pip.Based on the omics data and the validation results,the local resistance was analyzed by using the T-DNA insertion mutant sec5 of the At SEC.1 gene.The expression levels of resistance-related genes PR1 and PR5 were down-regulated,the count of bacterial colonies was increased,and the symptoms of leaf infection were aggravated,indicating that the Arabidopsis thaliana glucosyltransferase was induced by Pseudomonas syringae,and is related to plant disease resistance.In conclusion,At SEC.1、At SEC.2 and Fe SEC2 in Arabidopsis thaliana can affect the flowering transition,participate in the development of floral organs,and participate in the immune response to pathogens.These studies provide theoretical basis and new ideas for understanding SEC function in plant.