Study On The Regulatory Mechanism Of An AP2/ERF Transcription Factor,ThCRF1,in Response To Salt Stress In Tamarix Hispida

Ethylene-Responsive factors(ERFs)are plant-specific transcription factors(TF),which involve a variety of biological processes,especially in abiotic stress.However,ERFs of woody halophytes that are involved in salt stress are hardly studied.In this study,we studied ThCRF1,an ERF transcription factor that responds to salt stress from Tamarix hispida.This study systematically studied the mechanism of ThCRF1 regulation of salt tolerance.(1)ThCRF1 is a protein with a cDNA length of 1140 bp and encoding 379 amino acids.The bioinformatics analysis revealed that the ThCRF1 protein contained a typical AP2/ERF conserved domain and that there was a CRF conserved domain and a TEH interval at the N-terminus,so we named it ThCRF1.(2)Using the gene gun,yeast one hybrid(Y1H)and yeast two hybrid(Y2H)techniques,it was found that ThCRF1 protein is a nuclear localization protein with transcriptional activation domain which can be combined with cis-acting elements such as TTG,DRE and GCC-box.(3)The transient function of ThCRF1 transgenic T.hispida was used to analyze the function of the gene by gain and loss of function.It was found that ThCRF1overexpression(pROKII-ThCRF1-flag)in T.hispida significantly improved the tolerance of plants to salt stress,whereas the inhibitory expression of ThCRF1 gene(RNAi-ThCRF1)was significantly reduced the tolerance of plants to salt stress.Further qRT-PCR experiments showed that ThCRF1 induces the expression of genes including those encoding pyrroline-5-carboxylate synthetase(P5CS),trehalose-6-phosphate synthase(TPS),trehalose-6-phosphate phosphatase(TPP),superoxide dismutase(SOD)and peroxidase(POD),which lead to enhanced proline and trehalose levels and increased SOD and POD activities.The overexpressed ThCRF1 transgenic Arabidopsis thaliana was obtained by stable transformation,andthe gene function of ThCRF1 transgenic lines with relatively high expression level was studied by quantitative detection.The results further confirmed the results of T.hispida.(4)ThCRF1 transgenic A.thaliana line L2 was used to RNA sequencing and ChIP assay the results showed that ThCRF1 could bind to the promoter region of the regulated genes to induce the expression of these stress-related genes.In summary,ThCRF1 improves the tolerance of the plant to salt stress by enhancing the trehalose and proline biosynthesis to regulate the osmotic potential,increase the activity of SOD and POD to enhance the scavenging activity of active oxygen.