Abiotic Stress Tolerance Functions And Epitope Characteristics Of Arabidopsis Thaliana Telomerase Reverse Transcriptase

Telomerase,responsible for synthesis of telomere DNA in eukaryotic cell,is a ribonucleoprotein complex with reverse transcription activity.And telomerase reverse transcriptase(TERT)is the core component of telomerase.In the field of the TERT biology of animals and human,studies have shown that TERT has non-telomeric functions such as gene expression regulation,mitochondrial function regulation,stress protection,etc.However,the non-telomere function of plant telomerase reverse transcriptase remains elusive.Therefore,this study aimed to reveal the non-telomeric function of Arabidopsis thaliana telomerase reverse transcriptase(AtTERT)in Arabidopsis and E.coli.The main results are as follows:The eukaryotic expression vector pCAMBIA2300-GFP-AtTERT was constructed,and the subcellular localization of GFP-AtTERT protein was determined by transient expression of tobacco and stable expression of Arabidopsis thaliana.The results showed that the GFP-AtTERT fusion protein was mainly located in the nucleus,with only small amounts in the cytoplasm.The abiotic stress resistance of transgenic Arabidopsis thaliana showed that overexpression of AtTERT in Arabidopsis thaliana enhanced its salt tolerance,while no significant change in drought resistance was observed.pET32a/pGEX-4T-1-AtTERT was constructed as prokaryotic expression vector,then genetically transformed into E.coli and AtTERT protein expression was induced for functional analysis.The result showed that the salt tolerance,drought resistance and hydrogen peroxide tolerance were enhanced in E.coli transformed with AtTERT,while its freezing tolerance was reduced.Those results suggest that AtTERT plays an important role in the abiotic stress of Arabidopsis and E.coli.Based on AtTERT functional domain prediction,the prokaryotic expression vector pET32a-P1/2/3/4 with different functional domains of AtTERT was constructed,and the purified protein was induced,and then its reactivities were identified by ELISA.The effect of the conserved functional domain of AtTERT(P4)on response to abiotic stress was also explored.Protean software analysis showed that the AtTERT protein epitope was most likely located at the N-terminus and the middle of the amino acid peptide chain;The results showed that four proteins could react with TERT positive serum,while the antigenicity of P4 is strongest.And the salt tolerance,osmotic stress resistance were reduced in E.coli transformed with P4.Those results suggest that the integrity of AtTERT is the structural basis for its normal function.In summary,this study initially proved that AtTERT has non-telomere function,which lays a foundation for further research.