Fuctional Study Of The OsTRM13 Gene On Am Nucleoside Modification In TRNA

There are a large number of post-transcriptional modifications in tRNA,which are derived from the four basic nucleosides Uridine?Cytidine?Guanosine and Adenosine.The function of methylation modification and related methyltransferases have been extensively studied in yeast and human genes,but little has been done in plants.Therefore,in this paper,we tried to study the effects of tRNA methylation modification on rice growth and development and stress tolerance through the research methods of genetics,plant physiology and in vitro protein biochemistry.This paper mainly studied on the functions of rice OsTRM13 gene in Am nucleoside modifications.According to the results of previous studies in our lab:OsTRM13 gene can successfully replace Am modification in yeast Am deletion mutant strain,and in vitro prokaryotic OsTRM13 protein can catalyse tRNA^GlyGCC fourth base C/A to form Cm/Am modification,which demonstrat that OsTRM13 is similar to TRM13 in its ability to synthesize Am methylated nucleosides using tRNA substrates.Studies on the overexpressed material and RNAi material of OsTRM13 gene proved that the expression level of OsTRM13 gene affected the synthesis of Am nucleoside modification on tRNA and the normal growth of rice.The level of Am nucleosides modification in the overexpressed materials was significantly increased,and the tolerance to high salt stress was improved;The level of Am nucleosides modification in RNAi materials was significantly reduced,and the tolerance to high salt stress was decreased?Wang et al 2017?.This topic from previous studies of rice OsTRM13 CRISPR mutant materials,continue to conduct the OsTRM13 gene function research,including the CRISPR mutation type surface analysis and modified test Am nucleoside,tissue specific gene expression,rice endogenous tRNA as the substrate of methylation in vitro experiment,as well as OsTRM13 CRISPR materials compared with wild type transcriptome analysis.The main results are as follows:1.The OsTRM13 CRISPR mutant material was shorter than WT,And the seed setting rate was lower than that of the WT;2.The modification content of Am nucleoside in OsTRM13 CRISPR mutant was significantly lower than that of the WT;3.GUS staining results showed that OsTRM13 was expressed in heads and stem nodes at booting stage;4.OsTRM13 can be induced in vitro but can not methylation the rice tRNA^MetCAT and tRNA^AspGTC;5.Transcriptome analysis showed that the differentially expressed genes of OsTRM13CRISPR mutant and WT were mainly related to oxidative reduction,transcriptional regulation,amino acid and nucleic acid metabolism,which partly explained the mechanism of OsTRM13 on the hypersalt stress phenotype.