Research On The Molecular Mechanism Of The Responses Of The Endophyte Symbionts Of Achnatherum Inebrians To Low Phosphorus

Phosphorus is crucial for plant growth and crop yield.Although phosphorus is abundant in the Earth’s crust,it usually exists in a form that plants cannot absorb,greatly limiting the efficiency of phosphorus absorption by plants.Grassland is the most frequently affected natural ecosystem by human activities,and its habitat is very sensitive to today’s global change.With the continuous increase of global nitrogen deposition,the limitation of available phosphorus on grassland productivity is becoming increasingly significant.Although research has shown that the application of phosphorus fertilizer increases the effectiveness of phosphorus in soil and alleviates phosphorus limitations to some extent,long-term reliance on mineral phosphorus fertilizer not only triggers the depletion of phosphorus resources and the increase in the cost of phosphorus fertilizer,but also leads to a decline in soil quality,severe acidification,reduced microbial activity,causing soil acidification,water eutrophication,and other environmental problems,which is undoubtedly inefficient and unsustainable.Achnatherum inebrians is a perennial herbaceous plant of the Gramineae family,widely distributed in northwest China.It usually grows in a low nutrient environment and forms a mutually beneficial symbiotic relationship with Epichlo(?) gansuensis,which has strong stress resistance.Therefore,studying the physiological and molecular mechanisms of A.inebrians under low phosphorus conditions can provide important functional gene resources for selecting resistant varieties using endophytic fungi,and also provide theoretical basis for developing new forage germplasm with excellent phosphorus tolerance.The subjects were A.inebrians(E+)with E.gansuensis and A.inebrians(E-)without E.gansuensis in this study.The seedlings of A.inebrians were treated with different phosphorus concentrations(0.01 m M P and 0.5 m M P).Physiological methods were used to analyze the biomass,metal element content,amino acid content,and phosphorus utilization efficiency of E+and E-A.inebrians under different phosphorus concentrations.Combined with metabonomics and transcriptomics,the molecular mechanism of E.gansuensis regulating the response of A.inebrians to low phosphorus was analyzed.The main results are as follows:(1)Under low phosphorus conditions(0.01 m M P),endophytic fungi significantly increased the dry weight of leaves of A.inebrians,and the dry weight of E+ leaves of A.inebrians increased by 30.5% compared to E-leaves of A.inebrians.E.gansuensis had no significant impact on the dry weight of the roots of A.inebrians.Under low phosphorus conditions,the nitrogen content in the roots of E+and E-A.inebrians was significantly increased,while the phosphorus content in the leaves and roots of E+and E-A.inebrians was significantly reduced.Low phosphorus reduced the phosphorus utilization efficiency(PUE)and total PUE of the leaves and roots of E+ and E-A.inebrians,but E.gansuensis significantly increased the PUE and total PUE of the leaves of A.inebrians under low phosphorus conditions,and increased by68.5% and 45.9%,respectively,thereby improving the growth of A.inebrians under low phosphorus conditions.In addition,under low phosphorus conditions,E.gansuensis increased the potassium content in the leaves and roots of A.inebrians,decreased the magnesium and iron content in the leaves of A.inebrians,and changed the distribution of metal elements in A.inebrians.(2)GC-MS non-targeted metabolomics analysis revealed that E.gansuensis significantly influenced changes in the content of 13 metabolites(4 up-regulated and9 down-regulated)in leaves and 17(6 up-regulated and 11 down-regulated)metabolites in roots of A.inebrians under low phosphorus conditions.Besides,the metabolic pathways that were affected by E.gansuensis on leaves of A.inebrians under low phosphorus conditions were biosynthesis of secondary metabolites,ascorbate and aldehyde metabolism,pyruvate metabolism,valine,and leucine and isoleucine biosynthesis;while cyanine metabolism,tyrosine metabolism,arginine and proline metabolism,and tryptophan metabolism in roots of A.inebrians were probably the most relevant ones regulated by E.gansuensis under low phosphorus conditions.In addition,E.gansuensis up-regulated 14 amino acids and downregulated 4 amino acids in leaves of A.inebrians,and up-regulated 7 amino acids and down-regulated 4 amino acids in roots of A.inebrians under low phosphorus conditions In conclusion,the E.gansuensis may have reorganized amino acid metabolism in leaves and roots of A.inebrians to help it adapt to low phosphorus.(3)Transcriptomic analysis revealed that under low phosphorus conditions,E.gansuensis affected 1225 differentially expressed genes(529 up-regulated and 696down-regulated)in leaves and 1516 differentially expressed genes(873 up-regulated and 643 down-regulated)in roots of A.inebrians,which were mainly involved in amino acid metabolism,carbon metabolism,secondary metabolism,ion transport,transcriptional and translational regulation,phytohormone signaling and other biological processes.The KEGG pathway enrichment analysis showed that the differentially expressed genes up-regulated by E.gansuensis were mainly enriched in carbon metabolism,glycerophospholipid metabolism,pentose phosphate pathway,glycolysis/gluconeogenesis,and starch and sucrose metabolism in the leaves of A.inebrians.The differentially expressed genes up-regulated by E.gansuensis were mainly enriched in selenium compound metabolism,carbon fixation in photosynthetic organs,fructose and mannose metabolism in the roots of A.inebrians.In summary,the present study has preliminarily investigated the physiological,metabolic and molecular mechanisms underlying the response to low phosphorus in the symbiosis of A.inebrians-E.gansuensis.The differential metabolites,key differential metabolic pathways and key differentially expressed genes in response to low phosphorus were identified,and the physiological mechanism of low phosphorus adaptation in the symbiosis was also initially explored,laying a theoretical foundation for understanding the mechanism of low phosphorus tolerance in A.inebrians by E.gansuensis,and providing a scientific basis for breeding excellent forage grass varieties with low phosphorus tolerance.