| Low temperature is an important factor that affects the growth and development of organisms,leading to a decline in yield and quality.Arthrospira platensis is an important economical microalgae.Large-scale breeding base of A.platensis has large geographical spread in China.The use of runway-style open culture and greenhouse cultivation can be affected by low temperature stress in varying degrees,especially in early spring and late autumn.Therefore,it is of theoretical and practical value to study the adaptability of A.platensis to low temperature and improve its cold-resistant ability for the protection of A.platensis.In this paper,the effects of low temperature stress(15?)and zinc ion treatment(75?M)on the growth and physiological parameters of A.platensis were studied.At the same time,based on transcriptome sequencing,proteomics was used to explore the mechanism of A.platensis in response to low temperature stress and zinc ion improving its low temperature resistance.The main findings are as follows:(1)In this experiment,the Illumina double-end sequencing technology was used to sequence the transcriptome of A.platensis,and 104,979 unigene sequences were obtained.The 104,979 unigenes obtained were annotated to the UniProt database,among which 47,062(44.8%)unigenes were annotated;45,685(43.5%)unigenes were annotated to the GO database;47,075(44.8%)unigenes were annotated to the COG database;24,162(23.0%)unigenes were annotated to 321 metabolic pathways in the KEGG database.A complete transcriptome database of A.platensis was established.(2)A total of 943 differentially expressed proteins were identified by proteomics technique between the control group(30?)and low temperature group(15?).Based on GO and KEGG annotation classifications,these differentially expressed proteins were classified into carbohydrate metabolism,photosynthesis,amino acid metabolism,and translation.Then we combined these proteins with physiological responses to analyze the low temperature response mechanism of A.platensis.Low temperature stress produces excessive reactive oxygen species that activate the antioxidant system,enhances the ability of removing reactive oxygen,and slows damage of reactive oxygen to biofilms and macromolecules in the cells.The level of enzymes related to gluconeogenesis and polysaccharide degradation in carbohydrate and energy metabolism are up-regulated,and the level of enzymes involved in polysaccharide anabolism are down-regulated.This indicates that low temperature stress is beneficial to the accumulation of soluble sugar in A.platensis.Increased glutathione and glutamate-related enzymes in the amino acid anabolic pathw ay suggest that low temperature may up-regulate some specific amino acids(such as proline)involved in low temperature stress.The study also found that some increased stress defense proteins,such as RNA helicase,actively respond to low temperature stress.In addition,under low temperature stress.the level of antenna proteins,photoreaction center proteins and key enzymes in the Calvin cycle are down-regulated in A.platensis,so the photosynthesis rate is reduced.Translation-related enzymes level are also suppressed.Therefore,A.platensis positively responds to low temperature stress by up-regulating some stress defense proteins,and enzymes involved in soluble sugars and amino acid biosynthesis,as well as activating antioxidant systems.(3)Based on proteomics,the mechanism of zinc ion improving the low temperature resistance of A.platensis was studied.74 differentially expressed proteins were identified between the low temperature group and the zinc ion group(LT+zinc).Based on GO and KEGG annotation classifications,these differentially expressed proteins were classified into photosynthesis,carbon metabolism,nucleotide metabolism,translation and amino acid metabolism,and signal transduction pathways.In combination with physiological changes,the mechanism of zinc ion improving the low temperature resistance of A.platensis was analyzed.Under low temperature stress,after adding zinc ion,some antenna proteins,photoreactive center proteins and enzymes involved in the synthesis of pigments increased,which improved light absorption and electron transfer efficiency,then enhanced photoreaction.Some key enzymes in the carbon fixation pathways were also up-regulated,which increased photosynthetic carbon fixation rates.This suggests that overall photosynthesis increased with the addition of zinc ion.At the same time,the antioxidant system is further enhanced,which increased the ability of removing intracellular reactive oxygen species,and effectively reduced membrane lipid peroxidation and damage to macromolecular substances.Some of the enzymes involved in the synthesis of amino acids and nucleotides also increased,indicating that zinc ion can increase the rate of anabolic metabolism of amino acids and nucleotides and facilitate the growth of A.platensis.In addition,the levels of proteins(histidine kinase)which enhance signal transduction increased,indicating that zinc ion helps cell actively sense environment and responds to stress.Therefore,zinc ion increases the anabolic rate of A.platensis,which is conducive to positively respond to low temperature stress.In summary,high-throughput sequencing technology was used in this study to perform large-scale transcriptome sequencing of A.platensis.After annotating and analyzing the obtained genes,we constructed a transcriptional database of A.platensis.Based on the proteome level,the mechanism of A.platensis in response to low temperature stress was revealed,and a method for improving the low temperature resistance of A.platensis was explored.This study has an important significance both in theory and in practical application,and it also laid a solid foundation and provided important research clues to further study on the low temperature adaptation.It has opened up new ways for A.platensis breeding bases to increase yield and quality in low temperature stress,especially in early spring and late autumn.It is of great significance to prolong the breeding period and promote the development of A.platensis industry. |
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