The Biosynthesis And Roles Of Amino Acid Bioactive Substances In Regulating Tomato Growth And Development Under High Temperature And Drought Stress

Tomato is an important vegetable crop with abundant nutrition,wide cultivation area,and the highest yield among all vegetable crops.It prefers exist in warm temperature but not tolerant to heat during the cultivation process.Additionally,most tomato varieties are relatively tall,and belong to the continuous flowering and fruiting type with relatively large demand for water.With global warming and water shortage,extreme weather such as high temperature and drought are becoming more frequent,greatly impacting crop growth and development.Therefore,exploring the regulatory mechanisms of tomato growth and development under high temperature and drought stress is of great production significance.The amino acid bioactive substances pipecolic acid(Pip)and phytosulfokine(PSK)have been widely reported to play roles in plant growth,development,and stress responses,but their effects on plant growth and development under high temperature or drought conditions are still unclear.This study explored the biosynthesis and roles of Pip and PSK in regulating tomato growth and development under high temperature or drought stress.Genetic and transcriptomic approaches were used to reveal the regulatory mechanisms in-depth.The main findings are as follows:1.Regarding the unclear role of Pip in plant response to drought stress,the study investigated the response of Pip biosynthesis pathway to drought stress and its regulatory effects on tomato growth and development.It was found that Pip biosynthesis gene Sl ALD1 and hydroxylated gene Sl FMO1 responded to drought stress at the transcript level.Slald1 and Slfmo1 mutants were generated by CRISPR/Cas9 gene editing approach.Drought resistance dramatically increased in Slald1 mutants compared with wild-type(WT),which was associated with increased CO2 assimilation,photosystems activities,antioxidant enzymes activities,ascorbate and glutathione content,and reduced ROS accumulation,lipid peroxidation and protein oxidation.On the contrary,Slfmo1 mutants were more sensitive to drought,showing damaged photosystems and impaired antioxidant systems,which were significantly alleviated by exogenous ascorbate.2.Further study of the role of the Pip biosynthesis pathway in regulating tomato fruit ripening and quality formation revealed that both Slald1 and Slfmo1 mutant fruits were smaller than the WT.As the Slfmo1 mutant was overly sensitive to drought,we focused on observing fruit under normal conditions and found that the Pip biosynthesis pathway could simultaneously promote tomato ripening and improve fruit quality.Both the Slald1 and the Slfmo1 mutants exhibited delayed onset of ripening,decreased fruit size,and reduced nutrition and flavor.Exogenous treatment with Pip and NHP promoted fruit ripening and improved fruit quality.Transcriptomic analysis combined with weighted gene co-expression network analysis revealed that the genes involved in the biosynthesis of amino acids,carbon metabolism,photosynthesis,starch and sucrose metabolism,flavonoid biosynthesis,and plant hormone signal transduction were affected by Slfmo1 mutation.Transcription factor prediction analysis revealed that the NAC and AP2/ERF-ERF family members are notably involved in process regulation.3.Regarding the unclear role of PSK in tomato response to high temperature.PSK biosynthesis gene PSK3 and PSK3 L were found significantly responded to high temperature.Tomato PSK receptor pskr1 mutants were generated by CRISPR/Cas9 gene editing approach,it was found that the mutant was more sensitive to hightemperature than the WT,with significantly higher levels of membrane lipid peroxidation and electrolyte leakage.Further exploration of the regulatory mechanism revealed that the photosystem activity of pskr1 mutant was lower under high-temperature stress,and the level of accumulated ROS was significantly higher.4.Further exploring the response of PSK signaling in pollen germination under high temperature stress,it was found that pskr1 mutant showed weaker resistance to high temperature,with lower pollen vitality and germination rate.Flow cytometry was used to detect the ROS content of pollen,revealing weaker ROS signals in pskr1 mutant pollen under high temperature conditions.The interaction between PSKR1 and NADPH oxidase Rboh B was verified by Bi FC and Co-IP methods,and PSKR1 can phosphorylate Rboh B as a kinase.The study preliminary indicates that plants can establish a link between PSK signals and ROS signals in regulating pollen fertility under high temperature,through interactions between PSKR1 and Rboh B.In summary,this study explored the biosynthesis and roles of Pip and PSK in regulating tomato growth and development under high temperature and drought stress.Providing theoretical basis for cultivating green,safe,and high-resistance germplasm for agricultural production.