Physiological And Biochemical Mechanisms Of Exogenous 5-ALA Relieving Drought Stress In Wheat Seedlings

Drought stress is one of the most important multidimensional abiotic stress factors,which affects plant growth metabolism and reducing the final yield.5-aminolevulinic acid(5-ALA)is a physiologically active substance in plants,which has the potential to improve agricultural production.In this study,we use isotope-labeled relative and absolute quantitative proteomics(i TRAQ)to analyze the proteome levels of chloroplasts in wheat seedlings under drought stress by exogenous5-ALA,and use q RT-PCR to analyze the differentially expressed proteins at the gene level.Verification was performed to determine leaf relative water content(RWC),chlorophyll content,malondialdehyde(MDA)content,proline content,soluble protein content,ATPase content,NADPH enzyme content,and SOD,POD activity.To investigate the role of exogenous 5-ALA in relieving the protein regulatory network and physiological regulation of wheat seedlings under drought stress.Experiment choose Aikang 58 wheat varieties as experimental material,in the period of two leaves and a heart for spraying on the leaf concentration of 100 mg L-15-ALA in simulated natural dry after 6 d,the experiment was carried out on the leaf of the second wheat seedling..Experimental results show that the 5-ALA pretreatment significantly improved the wheat seedlings under drought stress the leaf relative water content(RWC),the content of Chlorophyll a and Chlorophyll b,maintain the normal growth of wheat seedlings,reduced the Malondialdehyde(MDA)content,at the same time increase the content of proline,soluble protein content,ATPase content,NADPH enzyme content and increase SOD and POD activity;A total of 2,442 proteins were identified by the i TRAQ analysis,and the related differentially expressed proteins were verified at the genetic level.The results showed that under drought stress,5-ALA could significantly improve the tolerance of wheat seedlings under drought stress.The main experimental results are as follows:1 Regulation of exogenous 5-ALA on the growth of wheat seedlings under drought stressThe growth of wheat seedlings was significantly inhibited under drought stress.With the continuous strengthening of the stress intensity,the damage to wheat seedlings became more severe,and cause death of wheat seedlings when the drought time reached 12 days.When the drought time reached 6 days,the relative water content(RWC)and chlorophyll content of wheat seedling leaves decreased significantly.When the drought time reached 9 days,the RWC and chlorophyll content of wheat seedling leaves decreased more significantly.The relative water content(RWC)and chlorophyll content of wheat seedling leaves decreased significantly at the 6th day of the drought period,and decreased much more significantly at the 9th day of drought.Meanwhile drought stress also exacerbated the membrane lipid peroxidation and malondialdehyde(MDA)content in wheat seedlings.Exogenous 5-ALA significantly increased the relative water content(RWC)and chlorophyll content of wheat seedling leaves under drought stress,relieved the increase of MDA content under drought stress,and enhanced the stability of cell membrane and promoted photosynthesis and other life activities to alleviate the drought stress on the growth of wheat seedlings.Thus holding the normal growth of wheat seedlings.2 The effect of exogenous 5-ALA on antioxidant system of wheat seedlings under drought stress.Drought stress can lead to wheat seedling excessive accumulation of reactive oxygen species in the body,and wheat seedlings by regulating the balance of antioxidant enzyme system response to stress.In this experiment,drought stress can induced significantly increased leaves superoxide dismutase(SOD),peroxidase(POD)in wheat seedling,but when the stress exceeds the regulation of plant antioxidant enzyme system of threshold,the balance of antioxidant enzyme system will be damaged.For example,POD activity decreased when drought stress reached 9d and6 d.Exogenous 5-ALA treatment can significantly improve the activity of SOD and POD under drought stress,it is the role of exogenous 5-ALA can enhance the ability of antioxidant enzyme system with ROS,and reduce the wheat seedling damage.3 Exogenous 5-ALA regulates ATPase and NADPH enzyme content in wheat seedlings under drought stressDrought stress will inhibit the photosynthetic system of wheat seedlings and reduce the photosynthetic rate,which leads to the decrease of photosynthetic system components ATPase and NADPH enzyme.Exogenous 5-ALA treatment significantly increased ATPase and NADPH enzyme levels,thereby maintaining efficient photosynthesis.4 Exogenous 5-ALA regulates proline content and soluble protein content in wheat seedlings under drought stressDrought stress induces plants to accumulate more infiltrating material to mediate the body's water potential,thereby maintaining normal metabolic activity.In this experiment,drought stress induced a significant increase in proline content of the osmoregulator,but significantly reduced the soluble protein content.The exogenous5-ALA treatment not only increased the soluble protein content,but also promoted the further increase of proline content.Therefore,the tolerance of wheat seedlings to drought stress was effectively improved.5 Isobaric tags for relative and absolute quantification(i TRAQ)analysis and verification of relevant differential proteins at the gene levelIn this experiment,2442 proteomes were identified through i TRAQ analysis of the sample chloroplasts.727 key differential proteins that were regulated with drought stress expression and exogenous 5-ALA were successfully screened through bioinformatics analysis.Related proteins such as photosynthetic synthesis-related,photosynthesis-related,energy metabolism-related,protein metabolism-related,amino acid synthesis-related,antioxidant-related,protein synthesis and transport-related proteins can be used as main targets for studying the mechanism of exogenous 5-ALA on the drought response mechanism of wheat seedlings.We selected 12 representative proteins(their search numbers in the uniprot.database were A0A1D6BFT6,A0A1D5YCF1,Q8S9H0,A0A1D5SMS3,Q43202,A0A1D6S578,A0A1D5WTV3,A0A1D5ZBT2,A0A096UTK7,A0A080YUW8,A0A1D6BSD3,A0A0C4BIL8)using q RT-PCR to verify their expression level under drought stress.q RT-PCR analysis showed that the gene expression patterns of these differential proteins were consistent with the results of bioinformatics analysis.