Response Analysis Of Sugar Metabolism To Salt Stress In Cassava(Manihot Esculenta Crantz)

Cassava(Manihol esculenta Crantz)is an important tropical cash crop,and it is an important source for food and bioenergy in the tropics.Salt stress,which causes osmotic stress and oxidative stress on plants,is the main environmental factor affecting the normal growth of plants and limiting crop yield.Sugars can be used as osmotic adjustment substances and signal molecules to participate in stress response.Sucrose is the main form for long-distance transport of sugars from source to sink.Sucrose Phosphate Phosphatase(SPP)is closely related to sucrose synthesis,while Sugar Transport Proteins(STP)play an important role in the transport and distribution of carbohydrates.Flavonoids can be involved in plant stress response as antioxidants,and their content is positively correlated with the activity of Phenylalanine ammonia-lyase(PAL).The effects of sugars and amino acids on the adaptation of cassava to salt stress were analyzed,and the response of MeSTPs,MeSPPs and MePALs to salt stress was analyzed,which could provide a theoretical basis for the subsequent selection and cultivation of cassava salt-tolerant varieties.In this study,cassava KU50(Kasetsart University 50)was used as experimental material.Firstly,the morphological and physiological changes of cassava under salt stress were analyzed.The contents of 17 amino acids and 8 carbohydrates were analyzed by high performance liquid chromatography(HPLC).Secondly,two gene family members of AleSPPs and MePALs were cloned by RT-PCR and analyzed by bioinformatics.Finally,the expression patterns of MeSTPs,MeSPPs and MePALs were analyzed based on transcriptome data and qRT-PCR experiments,and the most up-regulated member was selected from each family to be subjected to subcellular localization analysis by cassava protoplast transient expression system.The main results are as follows:(1)After 15 days of salt stress treatment(300 nM NaCI),the plant height and fresh weight of cassava decreased by 23.4%and 32.1%respectively.The number and size of leaves,the total number of roots and the number of lateral roots were significantly reduced.These results indicated that salt stress inhibited the growth and development of cassava’s leaves and roots,leading to the reduction of plant height and biomass.(2)Compared with the control,salt treatment increased the PAL activity by 61.2%and 49.2%in the leaves and roots respectively,and the flavonoid content by 64.5%and 61.6%in the leaves and roots respectively.However,the POD activity was very low in the leaves,and no significant change had been detected in roots.The SOD activity was decreased by 44.7%and 13.6%in leaves and roots,respectively.These results speculated that the enzymatic antioxidant system did not play a significant role in the response of cassava to salt stress.Cassava alleviated the salt-induced oxidative damages by improving non-enzymatic antioxidant capacity,specifically,by increasing PAL activity and promoting the synthesis of downstream flavonoids.(3)Compared with the control,salt stress increased the content of proline in leaves and roots by 116.7%and 71.6%,respectively.The total content of seven soluble sugars increased by 73.9%and 20.8%in leaves and roots,respectively.These results indicated that proline and soluble sugar were involved in the salt response of cassava.(4)Three members of the sucrose phosphate phosphatase gene family were cloned from cassava and named MeSPP1,MeSPP2 and MeSPP3,with the nucleotide sequence length of 1278 bp,1284 bp and 1275 bp,respectively.Six members of the phenylalanine ammonia lyase gene family were cloned and named MePAL1-MePAL6,with the nucleotide sequence length of 2148 bp,2112 bp,2139 bp,2051 bp,2133 bp and 2136 bp,respectively.(5)The expression patterns of MeSPPs,MePALs and MeSTPs under salt stress showed that MeSPP2 was up-regulated in cassava roots,and MeSPP3 was up-regulated in cassava leaves.In leaves,MeSTP2,MeSTP12,MeSTP16 and MeSTP20 were up-regulated.In roots,MeSTP7,MeSTP15 and MeSTP16 were also up-regulated.In MePALs family,only MePAL4 was up-regulated in roots,while other genes were down-regulated.The genes with the most up-regulated level in each gene family were MeSPP2,MePAL4 and MeSTP15.These results indicated that MeSPPs,MePALs and MeSTPs were involved in the salt response of cassava.(6)Subcellular localization analysis showed that MeSPP2 and MePAL4 were localized in the cytoplasm while MeSTP15 was localized on the cell membrane.