The Responses Of Ginkgo Biloba To Drought,salt And Heat Stresses

Abiotic stresses are the main environmental factors limiting plant growth and development,including drought,salt and heat.Plants can response to these stresses through their cytological,physiological,biochemical,molecular changes,and make responsive adjustments to adapt to the adverse environment.Ginkgo biloba has strong adaptation to the diverse environments.However,little is known about the mechanism of G.biloba responses to the main stress factors.In the present study,using G.biloba seedlings as experimental materials,we studied the cytologic,physiologic,molecular responses of G.biloba to drought,salt and heat stresses.The main results were as follows.（1）Through the observation of confocal laser microscopy,we found that the chloroplasts regularly distributed near the cell membrane of leaves in control.However,under drought,salt and heat treatment,chloroplasts showed irregular dispersive distribution.In addition,stomata exhibited different degree closures,especially under heat treatment.（2）Through physiological measurement,we found that ABA,proline,H2O2 concentration and the activity of CAT increased,and IAA concentration decreased in leaves under drought,salt and heat stress.However,the activity of SOD significantly increased in leaves under salt and heat stress,while no significant change under drought.Soluble sugar concentration increased significantly under drought and heat stress except for the salt stress.In addition,soluble protein concentration significantly increased under heat stress,but decreased under drought and salt stress（3）We further performed transcriptome sequencing,and screened 397,1007 and 4,494 differential genes from drought,salt and heat libraries respectively.Under drought stress,the differential expressed gene enriched in "flavonoid biosynthesis","photosynthesis","alanine,aspartate and glutamate metabolism","zeatin biosynthesis","plant hormone signal transduction".Under salt stress,the differentially expressed genes enriched in "flavonoid biosynthesis","zeatin biosynthesis","glycine,serine and threonine metabolism","plant hormone signal transduction Under heat stress,differential genes were enriched in "ribosome","arginine and proline metabolism","zeatin biosynthesis”.（4）In ABA metabolic pathway,under drought,salt and heat stress,three NCED genes Gb 41235,Gb₄1230 and Gb₀4986 were up-regulated,which related to synthesize xanthoxin in the ABA synthesis pathway.The AA03 gene Gb 05183 related to synthesize abscisate showed slightly up-regulated under heat,and no significant difference under both drought and salt stress.In ABA signal transduction pathway,most SnRK2 genes showed up-regulated expression under all three stress treatments.In addition,Gb₁1734,Gb₁6168,Gb₁6382 and Gb 11735 related to ABF were significantly up-regulated under heat stress（5）The association analyses between transcriptome and metabolome showed that the expressions of G6PDH（Gb₂6169,Gb₂6174,Gb₄1272）and 6PGDH（Gb 19030,Gb₁9037）in the pentose phosphate pathway（PPP）were down-regulated under heat stress.The genes（Gb 19030,Gb 19037,Gb₁7714）involved in the synthesis of D-Ribulose-5P and Ribose were also down-regulated under heat stress,and the contents of D-Ribulose-5P and Ribose decreased significantly.However,under drought and salt stress,G6PDH and 6PGDH showed no significant difference,and Gb 17714 involved in the synthesis of Ribose was up-regulated.（6）According to the analysis of amino acid metabolism,in glycine,serine and threonine metabolism,genes related to glycine synthesis were up-regulated under three stresses,including Gb 01536,Gb₀1539,Gb₃7290,Gb₂3485,Gb₀6259 and Gb₂7241.In addition,Gb₀3208 and Gb 35734 involved in glycine degradation pathway were down-regulated,and the contents of glycine and threonine were significantly increased.In arginine and proline metabolism,Gb 13947 and Gb 16297 involved in 4-aminobutyric acid synthesis were up-regulated under heat stress.Furthermore,the content of putrescine decreased and 4-aminobutyric acid content increased significantly.（7）Transcription factors（TFs）play a key role in the regulation of stress response.In our dataset,we found that 287 DEGs were putatively identified as TFs.Among them,NAC transcription factor genes Gb₁2203,Gb₃ 7720 and Gb₁2202 were only up-regulated under drought stress,but down-regulated or not changed under salt and heat stress.GRAS transcription factor gene Gb 22850 was only significantly up-regulated under salt stress,while GRAS transcription factor gene Gb₃9436 was significantly up-regulated under drought,salt and heat stress.HSF transcription factor gene Gb 15358 is homologous to HSFA1 and only significantly up-regulated under high temperature stress（8）Analysis of sRNA showed that the expression miR160a was up-regulated,while its target gene related to auxin response factor Gb₂5507（ARF18）was down-regulated under drought stress.Under salt stress,miR172a was up-regulated and its target gene Gb 30123（AP2-like protein）was down-regulated.Under heat stress,the genes related to leucine zipper protein Gb 02083（HOX32）was negatively regulated by miR166d and novel₂0.（9）We analyzed types and functions of resistance genes in G.biloba.A total of 237 resistance genes were associated with 40 TF families.There are 43 genes involved in a-bisabolene synthase,35 genes related to ginkgo lactone biosynthesis,55 genes related to ginkgo flavonoid biosynthesis,75 NBS-LRR class resistance genes,29 genes related to leucine rich repeat（LRR）receptor-like serine/threonine-protein kinase flagellin-sensitive 2（FLS2）and EF-Tu receptor（EFR）.