Transcriptome Analysis Of Pohlia Nutans Under Salt Stress And The Role Of 12-oxo-phytodienoic Acid Reductase In Plant Stress Resistance

The harsh environment has restricted the growth of land plants in the Antarctic continent.Moss and lichens are numerous and widely distributed in the coastal zone of the Antarctic continent and its adjacent islands,and are the main producers of the Antarctic terrestrial ecosystem.The growth of Antarctic moss is affected by many factors including temperature,wind,humidity and UV-B radiation.Since the moss grows mainly near the Antarctic coast,the salinity in the soil is also a major factor affecting its survival and reproduction.Antarctic moss has unique physiological and genetic characteristics that make it highly adaptable to the polar environment.In this study,the differential expression of the Antarctic moss Pohlia nutans under salt stress was analyzed by transcriptome sequencing.The effects of salt stress on the antioxidant pathways,jasmonic acid pathway,flavonoid content and the expression level of key genes in the synthetic pathway were analyzed.The expression pattern of 12-oxo-phytodienoic acid reductase(OPR)in different stresses,three Arabidopsis overexpressing pure lines of OPR gene were constructed.The specific results are as follows:(1)Transcriptome analysis of Pohlia nutans in response to high salt.High-throughput sequencing was used to analyze the transcriptome of A.Pohlia nutansunder high salt stress.After extracting total RNA,constructing libraries,filtering unqualified sequences,and assembly steps,a total of 285,789 transcripts and 209,838 unigenes were obtained from the samples of Antarctic moss.According to the obtained unigenes,GO annotations were performed to predict their function.A total of 120,579 unigenes were obtained by BLAST software and GO database for annotation information,and were divided into more than 40 categories,including growth,membrane,and antioxidant activity.Of them,15,997 unigenes were annotated into the biological process,and 590 unigenes were further classified as responsive to stimulation and antioxidant activity,some of which may be related to salt stress.KEGG pathway analysis showed that the most enriched pathway terms were ascorbate and aldarate metabolism,endocytosis,phenylpropanoid biosynthesis,flavonoid biosynthesis,regulation of autophagy,and plant hormone signal transduction.Gene expression levels of salt-treated and control samples were calculated by FPKM method to identify differentially expressed genes.Differentially expressed genes(DEGs)analysisshowed that 1,340 unigenes were significantly upregulated and 831 unigenes were significantly downregulated.(2)Antioxidant pathway and related differentially expressed genes of Pohlia nutans under salt stress.P.nutans has a strong tolerance to high salt stress..nutans has a strong tolerance to high salt stress.Gene differential expression analysis showed ABA/jasmonate pathway-related genes,antioxidant enzyme genes,and flavonoid biosynthesis-related genes were significantly upregulated after salt stress.The contents of malondialdehyde(MDA),ascorbate and flavonoid,and the activities of antioxidant enzymes were enhanced under salt stress.Exogenous application of ABA,MeJA or PA increased the salinity tolerance of P.nutans.In addition,ABA or MeJA treatment upregulated the gene expression of antioxidant enzymes and flavonoid biosynthesis-related enzymes.Therefore,we propose that antioxidant enzymes and flavonoids are two major antioxidant defense systems in P.nutans under salt stress,which protect cells and scavenge reactive oxygen species(ROS)when they encounter abiotic stress.(3)Functional analysis of 12-oxo-plant diene reductase OPR of P.nutans under different stresses.Under high salt stress,low temperature stress and UV-B radiation,most of the OPR genes in P.nutans were upregulated.Therefore,we proposed that OPRs in P.nutans participate in a variety of abiotic stresses,and speculated that they could play an important role in resisting external stress.In addition,the protein sequence alignment and phylogenetic analysis of the genes were used to analyze the role of the gene in plant stress resistance and to construct an Arabidopsis overexpression line.The experimental materials were provided for subsequent analysis of the Antarctic moss resistance system,and it benefit the study of the response mechanism of OPRs gene to abiotic stress factors.