Functional Analysis Of F3'H Gene In Plant Stress Response

The area of arable land in my country is small,and the adversity of land salinization has made food security more prominent.Therefore,identification of important stress tolerance genes for molecular design breeding is an important part of modern agricultural technology.On the other hand,the identification of important metabolic small molecules related to salt and drought resistance,the development of spectral,high-efficiency,and green stress-resistant conditioning agents for application in chemical agriculture is a new research direction in modern agriculture,which can not only avoids problems such as molecular breeding being limited to transformed varieties,and it is of great significance to solve my country's food security issues.The identification of important secondary metabolites related to stress tolerance is very important for understanding plant stress response mechanisms,crop genetic improvement and increasing stress yields,but so far there are few reports on secondary metabolites and their mechanisms that have application prospects.Flavonoids,as secondary metabolites related to adversity response,are significantly enriched under abiotic stresses such as salt and drought.It is generally believed that flavonoids act as antioxidants to remove excess ROS and improve the stress tolerance of plants.At present,there is a relatively in-depth understanding of plant stress response network,but there are few reports on the role and mechanism of secondary metabolism in stress response.Among flavonoids,flavonols have the highest activity,including quercetin and kaempferol,both of which have a dynamic balance in the body.This paper explored the changes in the dynamic balance of kaempferol and quercetin under adversity stress and the function of the flavanone 3-hydroxylase(F3'H)gene,the catalytic enzyme of the two dynamic balance.The main results are as follows:1.Adversity stress inhibits F3'H expression and shifts the dynamic balance to kaempferolSince the dynamic balance between kaempferol and quercetin is mainly controlled by F3'H,we initially analyzed the stress response mode of F3'H gene and the effect of stress on dynamic balance.The results showed that in Arabidopsis and wheat,the expression of F3'H gene was significantly down-regulated after NaCl,ABA and H2O2 treatment,while the content of kaempferol(K)and quercetin(Q)increased significantly,but the Q/K ratio showed a downward trend as a whole,which was consistent with the results of the expression analysis.The results showed that salt stress shifted the dynamic balance of kaempferol and quercetin to kaempferol by inhibiting the expression of the F3'H gene.2.The shift of homeostasis to quercetin weakens the tolerance to adversity stressIn order to explore the function of the F3'H gene in depth,we constructed an AtF3'H overexpressing Arabidopsis line.Under normal conditions,the Q/K ratio of the overexpression line has no significant change compared with the wild type,but the Q/K ratio was significantly increased under salt stress,indicating that AtF3'H overexpression shifts the dynamic balance between quercetin and kaempferol to quercetin.Under normal conditions,there is no significant difference in germination rate and growth status between wild-type and overexpression lines.Under different stress,the germination rate of the overexpression lines was significantly lower than that of the wild type,the growth inhibition of the above-ground part was more significant,and the fresh weight of the plant was smaller.The results indicate that the dynamic balance shift to quercetin caused by AtF3'H overexpression weakens the plant's tolerance to adversity stress.3.The shift of homeostasis to quercetin changes the ROS scavenging ability and affects the stress response signaling pathwayDAB staining analysis showed that the hydrogen peroxide(H2O2)content between wild-type and AtF3'H overexpression lines under normal conditions was similar.After NaCl and Mannitol treatment,H2O2 levels increased significantly,while the overexpression lines increased significantly.However,the H2O2 level of the overexpression line after ABA treatment was significantly lower than that of the wild type.The results indicate that the decrease in salt tolerance caused by the shift of dynamic balance to quercetin may be related to ROS.However,compared with the wild-type,the expression levels of some genes encoding ROS production and elimination related enzymes in the overexpression lines did not change significantly,indicating the complexity of the dynamic balance between kaempferol and quercetin in regulating ROS levels.In addition,we found that compared with the wild-type,the expression levels of some stress-response signaling pathway genes in the overexpression lines changed,indicating that the dynamic balance between kaempferol and quercetin may affect the salt tolerance ability due to the ABA signaling pathway.4.Treatment with exogenous quercetin has a similar effect to the shift of dynamic equilibrium to quercetinThe results of pharmacological analysis showed that under normal conditions,the addition of quercetin had no significant effect on the germination rate of Arabidopsis and wheat.Under salt stress,high-concentration quercetin co-treatment enhanced the inhibitory effect on seed germination.As the concentration increases,the inhibitory effect gradually increases.At the same time,quercetin also enhances the inhibitory effect of salt stress on the growth of seedlings,which is related to the concentration.Exogenous quercetin treatment increased the excessive accumulation of H2O2 in the seedlings.These results are consistent with the shift of dynamic balance to quercetin caused by F3'H overexpression,and further indicate that changes in dynamic balance can affect the salt tolerance of plants.