Response Of Phenolics In Juniperus Rigida To Stress And Mechanism Of Adaptation

The morphology,physiology and metabolism of plants will undergo a series of changes to adapt to the environment and improve survival competitiveness under stress conditions.Among them,plant secondary metabolites,especially phenolics,play an important role in improving plant protection,resisting and adapting to stress,and coordinating with the environment.Their production and changes are more closely related to the environment than primary metabolites.Juniperus rigida is a key protected plant in the Loess Plateau region and contains many phenolic substances.Exploring the response of its phenolic substances to adversity and its adaptation mechanism is very important for theoretical research and production application of J.rigida.Therefore,two experimental models were adopted in this study.One of them was to study the response of J.rigida phenolics to altitude in the arid environment of Helan Mountain and the humid climate of Changbai Mountain.The other was to study the effects of drought stress and UV-B radiation stress on J.rigida phenolics in potted J.rigida seedlings,and analyze the phenolics of J.rigida from plant morphology and physiology.The adaptation mechanism of J.rigida phenolics to stress from the molecular level.The main results are as follows:(1)The response of J.rigida phenolics to altitude in Helan Mountain and Changbai Mountain was revealed.(1)The phenolics(total phenols,total flavonoids,podophyllotoxin,quercetin,rutin,hyperoside,kaempferin,and p-coumaric acid content)and antioxidant activity of J.rigida in Helan Mountain were higher than Changbai Mountain,which was related to the xerophytic structure and high CAT and PPO activity and MDA content of J.rigida in Helan Mountain.The phenolics and biological activities(antioxidant and antibacterial activity)of J.rigida at high altitude(2182-2340 m)in Helan Mountain were higher than that at low altitude(1568-1883 m),which were related to the obvious xerophytic structure of J.rigida at high altitude and physiological characteristics.The physiological characteristics including higher MDA content,lower antioxidant enzyme activities(POD,CAT,SOD,PPO)and photosynthetic pigments(Chla,Chlb,Car).The phenolics and biological activities of J.rigida at high altitude(1073-1203 m)in Changbai Mountain were higher than those at low altitude(838-885 m),which was related to the thickened of J.rigida cuticle,epidermis and hypodermis,high POD,CAT and SOD activities,high MDA content,and low photosynthetic pigment content.(2)The soil water content,soil organic matter,soil alkali hydrolyzable nitrogen,soil available phosphorus and soil available potassium at the high altitude(2182-2340 m)of Helan Mountain were 56.94%,59.60 g/kg,200.87 mg/kg,6.62 mg/kg and 259.78 mg/kg,respectively,which were equivalent to the soil fertility in the middle-high altitudes of Changbai Mountain,indicating that the high altitude soil fertility of Helan Mountain and Changbai Mountain was suitable for the synthesis of J.rigida phenolics,and the maintain the normal growth of J.rigida.The physiology,phenolics and biological activity of J.rigida in Helan Mountain and Changbai Mountain were mainly affected by altitude,soil organic matter,soil water content,p H and total phosphorus.(2)The response of J.rigida phenolics to drought stress was elucidated.(1)The phenolics such as quercetin,amentoflavone,rutin,hyperoside,kaempferol and formononetin had the lowest content,which was related to increase POD and PPO that could degrade phenolics under moderate drought stress(40-50%of saturated soil water content).Under severe(20-30%of saturated soil water content)or extreme drought(8-10%of saturated soil water content)stress,the phenolics such as podophyllotoxin,quercrtrin,amentoflavone,rutin,hyperoside,kaempferol and formononetin had the highest content,and the best antioxidant and antibacterial activities.The hyperoside,quercetin and formononetin play a major role in the antioxidant activity of phenolics.Podophyllotoxin,quercetin,amentoflavone,formononetin and kaempferol increase the antibacterial activity against Salmonella enteritidis and Klebsiella pneumoniae.(2)The histochemical localization of phenolics in J.rigida needles revealed that phenolics were significantly higher in sheath cells,secretory cells,spongy tissues,palisade tissue,epidermis,sclerenchyma and parenchyma,while xylem,phloem,sclerenchyma and hypodermis had almost no phenolics distribution.Under drought stress,the resin ducts were enlarged,the fenestrated tissues were tightly arranged while the spongy tissues were loosely arranged,and provided the sites for the synthesis and accumulation of phenolics.(3)From the plant physiological point of view,the increase of POD and CAT activity was unfavorable to the accumulation of total phenols and total flavonoids.The increase of CAT activity was favorable to the accumulation of quercetin,amentoflavone,formononetin and kaempferol.The enhanced photosynthetic capacity of the plants was favorable to the accumulation of total phenols and unfavorable to the synthesis and accumulation of quercetin.(3)The response of UV-B radiation stress on J.rigida phenolics was clarified.(1)Under C1(+0.8μW/cm~2UV-B)radiation stress,the content of total phenols is 3.38 mmol/g,podophyllotoxin,amentoflavone,hyperoside,kaempferol and formononetin were reached largest value,which were associated with the elevated antioxidant enzyme activities and photosynthetic pigments.With the increase of UV-B radiation stress,the activity of phenolics and antioxidant enzymes decrease,and the increase of photosynthetic pigments was gentle,indicating that low-intensity UV-B radiation improves the photosynthesis and accelerates the synthesis of defensive substances.Increasing the intensity of UV-B radiation damaged the needle cells of J.rigida,resulting in the destruction of antioxidant enzyme system,photosynthetic system,and phenolic substance anabolism.The synthesis of phenolics was reduced.The J.rigida needles treated with C3(+2.5μW/cm~2UV-B)intensity had the highest antioxidant activity,and the J.rigida needles treated with C2(+1.6μW/cm~2UV-B)or C3 had better antibacterial activity.Among the phenolics of J.rigida,rutin and quercetin play a major role in its antioxidant activity,which the content of rutin and quercitrin increased,the antioxidant activity improved.The increase of podophyllotoxin and quercetin content improved the antibacterial activity of J.rigida against Bacillus subtilis and Listeria monocytogenes.(2)After UV-B radiation stress,phenolics were significantly higher in sheath cells,secretory cells,sponge tissue,palisade tissue,epidermis,sclerenchyma and parenchyma than in control,while xylem,phloem,sclerenchyma and hypodermis were almost absent.After UV-B radiation stress,the mesophyll cells in the needles of J.rigida were all smaller than those in the control group.However,the nucleus of these cells were significantly larger,and the number of stomata increased,mostly in a closed state.The lower stomatal compartment was smaller,and the stratum corneum was thickened.(4)Transcriptome sequencing analysis of J.rigida seedlings treated with significantly different UV-B radiation assembled a total of 95248 Unigenes,of which 27655 Unigenes obtained annotation results.Under UV-B radiation stress,37 differentially expressed genes were identified to be involved in the synthesis of phenolics in J.rigida,mostly with significant positive correlations,mainly through the pathways of phenylpropane biosynthesis,flavonoid biosynthesis,and flavonol biosynthesis.Among the differentially expressed genes,4CL,CHS,CHI,FLS,and PLR were the key genes for phenolic synthesis.In summary,the synthesis and accumulation of phenolic compounds in J.rigida were not only affected by environment conditions such as altitude,drought stress and UV-B radiation stress,but also related to anatomical structure,antioxidant enzyme activity and internal factors of photosynthesis.This study provides an important basis for the study of plant response and adaptation mechanism to stress,and it has reference value for vegetation restoration and sustainable management of J.rigida forests in J.rigida distribution areas.