Comparative Transcriptomic,Proteomic And Metabolomic Profiling Of Symbiotic And Aposymbiotic Strains Of Hydra Sinensis

Some photosynthetic unicellular algae live in symbiosis with invertebrate cells,so animal host has obvious competitive advantages when food is scarce.The symbiont consisting of animal and unicellular algae is a complex system with high scientific value,related studies have focused on the symbiotic mechanisms such as compatibility,mutual control and the exchange of substance and energy between host and symbiotic algae.Our research group had prepared aposymbiotic strain of Hydra sinensis based on symbiotic strain of H.sinensis.On this basis,comparative transcriptomic,proteomic and metabolomic profiling of symbiotic and aposymbiotic strains of H.sinensis were studied in order to explore the responses of H.sinensis to symbiotic algae on the molecular level.The main results are summarized as follows:1.Comparative transcriptomic profiling of symbiotic and aposymbiotic strains of H.sinensis47.73 Gb Clean Data were obtained by transcriptome sequencing,and there were 68,504 assembled unigenes,14,508 of which were more than 1 kb in length.The results showed that there were 439 differentially expressed genes between symbiotic and aposymbiotic strains of H.sinensis,among which included 379 up-regulated genes and 60 down-regulated genes.A total of 45 KEGG metabolic pathways were enriched based on 439 differentially expressed genes.There were 9 metabolic pathways closely related to glycometabolism,among which are 4(glycolysis and gluconeogenesis,oxidative phosphorylation,carbon metabolism,and fructose and mannose metabolism)in the top 10 of all pathways according to enrichment significance.Furthermore,5 aliphatic acid metabolic pathways,6 pathways involved in nitrogen metabolism and two antioxidative pathways(lipoic acid metabolism ranking seventh in terms of enrichment significance among all enriched pathways and peroxisome)were also enriched.2.Comparative proteomic profiling of symbiotic and aposymbiotic strains of H.sinensis152 differentially expressed protein spots were screened out between symbiotic and aposymbiotic strains of H.sinensis by two-dimensional electrophoresis,and these protein spots were analyzed by Matrix-Assisted Laser Desorption/ Ionization Time of Flight Mass Spectrometry(MALDI-TOF-MS)after enzymolysis.There were 131 differentially expressed protein spots identified successfully by searching Swiss-Prot and NCBInr databases based on peptide fingerprint analysis,2 of which failed in function annotation.After merging the proteins with the same annotation function,90 valid differentially expressed proteins(among which included 47up-regulated proteins and 43 down-regulated proteins)were finally obtained.30 KEGG metabolic pathways were enriched based on these differentially expressed proteins,and there are mainly two types of pathways in the top 10 of all pathways according to enrichment significance.One type is the NOD-like receptor signal pathway involved in the biological immune system,the other type is four metabolic pathways(inositol phosphate metabolism,carbon metabolism,citric acid cycle,and fructose and mannose metabolism)that participated directly in glycometabolism.In addition,there are two glycometabolism pathways(glycolysis and gluconeogenic pathway and oxidative phosphorylation pathway),sphingolipid metabolism pathway associated with fatty acid metabolism and two amino acid metabolism pathways after the top 10 of all pathways according to enrichment significance.In addition,the result of protein interaction analysis showed the significant interaction between inositol phosphate metabolism,carbon metabolism,citric acid cycle and fructose and mannose metabolism,which participated in glycometabolism,and these four glycometabolism pathways were clustered directly in the metabolic network.3.Comparative metabolomic profiling of symbiotic and aposymbiotic strains of H.sinensisA total of 1572 features were detected using gas chromatography-mass spectrometry(GC-MS)technology,and 459 substances were identified.Subsequently,207 differential metabolites between symbiotic and aposymbiotic strains of H.sinensis were identified based on orthogonal partialleast squares analysis(OPLS-DA).After mapping the differential metabolites to the KEGG database,44 metabolic pathways involving 88 of 207 all differential metabolites were enriched.Among these enriched pathways,there are four types of metabolic pathways are noteworthy.Above all,10 differential metabolites were involved in the ABC(ATP binding cassette)transporter system that transport many kinds of substances across membranes,and this pathway ranks fifth in terms of enrichment significance among all enriched pathways.Then,there are 22 metabolites involved in 10 pathways closely related to glycometabolism,8 metabolites involved in 5 metabolic pathways related to fatty acid metabolism,and 56 metabolites involved in 11 nitrogen metabolic pathways.4.A comprehensive analysis based on three comparative omics studies has revealed the molecular responses of H.sinensis to symbiotic algaeIn the study,these results of KEGG metabolic pathway enrichment analyses based on differentially expressed genes,proteins and metabolites between symbiotic and aposymbiotic strains of H.sinensis have revealed five types of molecular responses of H.sinensis to symbiotic algae:(1)Both glycometabolism and fatty acid metabolism pathways were enriched based on the three omics data.The carbohydrates synthesized through photosynthesis by symbiotic algae can not only meet their own nutrient requirement but also be passed to host cells as a nutrient supplement.Previous research has shown that nutrient passed from symbiotic algae to host cell possibly in the form of carbohydrate,it might be the reason why glycometabolism pathways in symbiotic strain of H.sinensis became more active than that of aposymbiotic strain.In addition,active fatty acid metabolism pathways in symbiotic strain of H.sinensis indicates that lipids may also be an alternative carriers for nutrient transfer from symbiotic algae to hydra cells,and it is worthy of further investigation.(2)Nitrogen metabolism pathways were enriched based on the three omics data,and it could be associated with the fact that symbiotic algae live inside host cells and their nutrients must be obtained from host cells.More active nitrogen metabolism pathways in hydra cells should contribute to further transforming nitrogenous waste products emitted originally outside of the cell into some nitrogenous molecules or ions that can penetrate the membrane of symbiotic vacuole,because symbiotic algae need continuous nitrogen supply asplant.(3)Symbiotic algae are not directly immersed in the cytoplasm of host cell,but they are wrapped by a layer of host cell membrane outside their cell walls.ABC(ATP binding cassette)transporter system enriched based on the metabolomic analysis should be responsible for transporting nutrient substances across membrane between host cell and symbiotic algae.(4)As the host of symbiotic algae,hydra cells have been confronted with a problem – the level of reactive oxygen species in host cell could be raising because of the photosynthesis of symbiotic algae.The transcriptome analysis revealed a relationship between more active antioxidant metabolism pathway of symbiotic strain of H.sinensis and the elevated level of reactive oxygen species.(5)The proteome analysis showed the decrease of immunity level in symbiotic strain of H.sinensis,and it reflects the possibility that hydra cells down-regulate their immunity levels in order to tolerate and contain symbiotic algae.