Study On The Mechanism Of Spermidine Biosynthesis Gene SAMDC On Salt-alkali Resistance Of Tomat

The tomato variety’Solanum lycopersicum lycopersicum cv.Zhongshu No.4’has strong growth potential,moderate sweet and sour,round and positive fruit type,wide adaptability and high yield.It has strong resistance to tobacco Mosaic virus and Phytophthora infestans,but its salt tolerance is poor,and it is susceptible to salt and alkali stress in the growth process.It is of great significance to explore the mechanism of resistance to saline-alkali stress for maintaining the good character and optimizing the quality of’Zhongshu No.4’variety.Previous studies of our research group showed that exogenous Spermidine（Spd）could alleviate the damage of saline-alkali stress on tomato plants.S-adenosylmethionine decarboxylase（SAMDC）is a key rate-limiting enzyme in spermidine biosynthesis and is involved in various physiological reactions under stress.SAMDC can respond to drought,high temperature,low temperature,saline-alkali stress and other abiotic stresses.However,the mechanism of SAMDC in response to and regulation of saline-alkali stress in tomato crops remains unclear.Therefore,in this study,tomato was used as test material to explore the function of SAMDC under saline-alkali stress.The main research results are as follows:（1）Six SlSAMDC family members were identified from tomato genome.Local BLAST and hidden Markov statistical models were used to detect SAMDC members in the whole tomato genome,which were divided into two subfamilies according to their evolutionary relationships.Chromosome localization results showed that 6 SlSAMDC members were located on 4 different chromosomes.The results of domain and conserved motif showed that there were obvious structural differences among different subfamilies.The results of promoter cis-acting elements analysis showed that there were a large number of elements related to light,hormone（gibberellin,salicylic acid,abscisic acid,methyl jasmonate）induction and abiotic stress（drought,low temperature,anaerobic）induction and protein metabolism in the promoter region of SlSAMDC family members.The results of intra-species collinearity analysis showed that there was a pair of SlSAMDC homologous genes in tomato.The results of collinearity analysis between species showed that tomato SlSAMDC had collinearity relationship with Arabidopsis At SlSAMDC,melon Cm SAMDC,capsicum Ca SAMDC and poplar Pt SAMDC.（2）A SlSAMDC gene was cloned from the tomato variety’Zhongshu No.4’in response to abiotic stress and hormone induction.A SlSAMDC gene was obtained by PCR amplification.CDS（Coding sequence）length was 1083 bp,encoding 361 amino acid residues.The tomato plants were treated with abiotic stress,and the expression of SlSAMDC was significantly inhibited on the darkness treatment.SlSAMDC responds to salt stress,abscisic acid induction and auxin induction.（3）SlSAMDC overexpression and CRISPR/Cas9-mediated gene editing vectors were constructed,and the SlSAMDC expression levels of eight overexpressed lines（SlSAMDC-OE）were 2.25-5.57 times of those of WT under normal growth conditions,respectively.The five mutant lines（slsamdc-ko）obtained showed different base site mutations at two mutation targets,including substitution,insertion and deletion of base mutations,and SlSAMDC was basically not expressed under normal growth conditions.（4）Overexpression of SlSAMDC promoted the growth of tomato plants.The overexpression of SlSAMDC significantly increased the net photosynthetic rate and chlorophyll content of tomato plants,and reduced the water loss rate of leaves.The mutant SlSAMDC reduced the net photosynthetic rate and chlorophyll content of tomato plants,and increased the water loss rate of leaves.（5）Overexpression of SlSAMDC improved the osmotic regulation ability of tomato plants under saline-alkali stress.With the increase of saline-alkali treatment time,the content of osmotic regulator proline（PRO）in SlSAMDC-OE plants was significantly increased compared with WT plants,while the content of malondialdehyde（MDA）was significantly decreased compared with WT plants,while the MDA content of slsamdc-ko plants was significantly increased compared with that of WT plants,and the content of PRO was significantly reduced compared with that of WT plants.（6）Mutant SlSAMDC increased the level of reactive oxygen species in mutant tomato plants under saline-alkali stress.Under saline-alkali stress,more H₂O₂,O₂^·-were accumulated in slsamdc-ko plants than in SlSAMDC-OE and WT plants,and the peroxidase content was significantly reduced compared with that in SlSAMDC-OE and WT plants.（7）SlSAMDC regulates the salinity resistance of tomato plants by regulating osmoregulatory substances and peroxidase content.