Effects Of Simulated Acid Rain On Some Physiological Characteristics And Gene Expression Profiles Of Arabidopsis Thaliana

Acid rain is a serious worldwide environmental problem.It leads to acidification of environment and impacts the growth and development of plants.To provide an essential scientific basis for the identification of plant genes involved in acid-tolerance as well as to genetically engineering crops and forest plants tolerant to acid rain,in this thesis study I have analyzed the physiological characteristics and gene expression profiles of Arabidopsis thaliana under the treatment of simulated-acid rain(SiAR).Four different of pH values,pH 2.5,3.0,4.0,5.6(control) in a gradient were chosen to study the effects of SiAR on the physiological properties of wide-type Arabidopsis(WT) and the Ca²⁺-sensing receptor(CAS) antisense lines(CASas).The results showed that,one week after SiAR treatment,the content of chl a/b and the activities of SOD and CAT were reduced along with the pH value of SiAR in both WT and CASas plants.Apparently,the area of leaf necrosis,soluble protein content,MDA content and POD activity were increased.In comparison,overall,pH < 3.0 SiAR would cause serious damage to the Arabidopsis thaliana,and WT was more tolerant to SiAR than CASas.In order to further explore the mechanism of plant response to SiAR,Affymetrix GeneChip ATH1 arrays were used to study the gene expression profiles of WT under pH3.0 SiAR treatment at 8 h,32 h,68 h and 116 h(Sample numbers:A8,A32,A68, A116,C8,C32,C68,C116).The expression levels of 439 genes were changed significantly and among these genes 394 had functional annotations or putative annotations in the public databases.They can be classified into 15 categories and are involved in several biological processes,such as metabolisms,interaction with the environment,and signal transduction.After 8 h,32 h,68 h and 116 h SiAR treatment,the transcripts of 55,66,25,and 30 genes were significantly changed compared to their corresponding controls, respectively.Among them,28,24,19 and 14 genes were induced,27,42,6 and 16 genes were repressed,respectively.Comparison between samples of the treatment, number of differential expressed genes of A8 and A32 is 35,including 15 up-regulated genes and 20 down-regulated genes.The gene expression difference between A32 and A68 is extremely significant.The number of differential expressed genes were 370,which contains 203 up-regulated genes and 167 down-regulated genes.We conjecture that there is a tolerant threshold value of Arabidopsis to pH 3.0 SiAR after treatment for 32 h to 68 h,when various physiological responses changed dramatically and expression levels of gene showed significant differences.Through the comprehensive analysis of gene expression levels,we have found that the transcription of 13 genes encoding calcium binding proteins and 15 genes associated with active oxygen metabolism was altered significantly,which indicates calcium signal transduction and active oxygen metabolism play important roles in plant response to SiAR.The expression levels of genes encoding zinc finger transcription factor protein,such as NAC transcription factor,ERF/AP2 transcription factors and MYB transcription factors,were in rising trend along with the increase in time and intensity of SiAR treatment.Acid tolerance may be improved by regulating the expression levels of these transcriptional genes.In addition,the expression levels of 37 genes associated with secondary metabolisms were changed significantly. Secondary metabolites,such as lignin and alkaloids products,were increased,which could improve the acid resistance of plants.Seven genes encoding light-harvesting chlorophyll proteins were up-regulated to maintain the activity of photosynthesis.It seems that in response to plants use a complex network involving multi-genes and in a multi-channel mode.The altered expression levels of these genes at the genomic level indicate that Arabidopsis employs a variety of ways to fight against acid rain.We have also discovered that genes in the salicylic acid(SA) pathway were up-regulated at the beginning of the acid rain treatment,but down-regulated when the plants were exposed to SiAR for 68 h.Interestingly,the genes in the jasmonic acid (JA) pathway were substantially up-regulated in response to acid rain.Taken together, we conclude that SiAR activates the SA pathway at the beginning,while the JA pathway gradually takes over with the increase in time and intensity of the acid rain treatment.By means of bioinformatics tools,we have also comprehensively analyzed how 7 different abiotic stresses,including SiAR,low temperature,drought,osmotic stress, wounding stress,genotoxic stress and UV-B,affect the gene expression in Arabidopsis thaliana at the genomic level.Our results show that the Arabidopsis gene expression profile for SiAR is most similar to that for wounding stress,followed by drought and genotoxic stress,and displays the minimum relevance to UV-B and osmotic stress.