Study On The Salt Tolerance Mechanisms Of Ion Compartmentation In Halophyte Halogeton Glomeratus

Soil salinization has become a global concern for the world,and it covers more than100 countries.According to the statistics of UNESCO and FAO,the salt-affected soils is approaching 1 billion hectares,with 50%of the all irrigated land around the world suffering from the soil salinization,and the amount is rapidly increasing at the speed of 10 million hectares a year.In china,the salt-affected cultivated land is more than 36 million hectares.A total of 250 thousand hectares arable land has to be abandoned due to soil salinity each year.The decrease of the grain yields caused by soil salinization is up to 20.7 billion kg each year.In 21 century,soil salinization will also bring a series of environmental problems,which affect the sustainable development of agriculture and ultimately affect human life.The key to solve the problem of soil salinization is to study the mechanisms of salt tolerance,explore salt tolerant genes and cultivate new salt tolerant crop varieties.Halogeton glomeratus(H.glomeratus)is an extreme halophyte that is widely distributed in arid regions of Northwest China,Mongolia and the Central Asia.It grows well on the desert and salt soil areas,thus not only having tolerance to salt,but also drought.This species is an ideal wild resource for obtaining salt tolerance and drought resistance genes.However,little is known about the adaptive mechanism of tolerance to salt in this species.In the present study,we systematically analysed the salt tolerance mechanisms of H.glomeratus by morphology,physiology,biochemistry,cytology,transcriptome and proteomics methods.The main findings are as follows:1.H.glomeratus is a typical halophyte belonging to the category of tolerance of tissue to accumulate Na~+or Cl~-,and it showed optimal growth under 100 mM NaCl condition.The primary physiological strategies to response salt stress in seedlings of H.glomeratus are to maintain high tissue water content(TWC),stomatal aperture,and adjust ion balance.2.The distribution of Na~+in seedlings roots,stems,and leaves is obviously different,and the highest content in leaves,followed by stems,and roots are lowest.The succulent leaves tissue is the main organ of salt enrichment.Furthermore,Na~+is compartmentalized predominantly into the leaf water-storage tissues,and finally it is compartmentalized into the cell vacuoles of water-storage tissues.3.Illumina RNA-sequencing was performed in five sequencing libraries that were prepared from leaf samples treated with 200 mM NaCl for 6,12,24 h and 72 h and a control sample to investigate changes in the H.glomeratus transcriptome in response to salt stress.The de novo assembly of five transcriptomes identified 50,267 transcripts.Among these transcripts,31,496(62.66%)were annotated,including 44 Gene Ontology(GO)terms and128 Kyoto Encyclopedia of Genes and Genomes(KEGG)pathways.One hundred and eighteen salt-induced genes were common to at least two stages of salt stress,and 291up-regulated genes were common to various stages of salt stress.Numerous genes that are related to ion transport,reactive oxygen species scavenging,energy metabolism,hormone-response pathways,and responses to biotic and abiotic stress appear to play a significant role in adaptation to salinity conditions in this species.At the same time,it is necessary to verify whether these genes of unknown function play important roles in response to salt stress in the further study.4.A combination of physiological characteristics and iTRAQ-based proteomic approaches was conducted to investigate the molecular mechanisms underlying the salt response of suspension cell cultures of halophytic Halogeton glomeratus.These cells showed halophytic growth responses comparable to those of the whole plant.In total,87up-regulated proteins and 192 down-regulated proteins were identified as common to both200 and 400 mM NaCl concentration treatments.Such salinity responsive proteins were mainly involved in energy,carbohydrate metabolism,stress defense,protein metabolism,signal transduction,cell growth,and cytoskeleton metabolism.5.To more fully characterize the salt stress response,we firstly employed Pacific Biosciences long-read sequencing technology for sequencing of the H.glomeratus transcriptome and identified 54,835 consensus isoforms with a mean length of 2,663 bp.Subsequent RNA sequencing(RNA-Seq)of leaves from plants subjected to 0,100,200,or400 mM NaCl concentrations was carried out,and a total of 115 upregulated and 87downregulated differentially expressed isoforms(core DEIs)were identified in samples from plants subjected to various levels of salt stress.In addition,a total of 16 upregulated core DEIs of unknown function were predicted to possess transmembrane domains(TMD),suggesting that these candidate isoforms could be involved in Na~+transport in H.glomeratus.6.As mentioned above transcriptome and proteomics analysis of H.glomeratus response to salinity stress,the known genes(proteins)regulating Na~+compartmentalization into the vacuole(NHX)and its efflux from the cytoplasm(SOS1)did not change significantly under salinity stress.As detailed in our findings above,the salt tolerance mechanisms of ion compartmentation in H.glomeratus have been preliminarily revealed.But so far we were unable to clearly address the question of how sodium is compartmentalized into the vacuoles and efflux from the cytoplasm of H.glomeratus,we speculated that different sodium transporters may be present in H.glomeratus and may efficiently compartmentalize sodium in the vacuole and prevent its accumulation in the cytoplasm.Certainly,it is necessary to the further study to characterize sodium transporters in H.glomeratus.