Molecular Mechanism Of Low Temperature Induced Synthesis Of Polyphenols In Tobacco Leaves At Seedling Stage

Low temperature is a kind of natural disaster often encountered in crop cultivation and an important factor restricting agricultural production in many areas.Under low temperature stress,many metabolic processes of plants,such as photosynthesis,carbohydrate metabolism,lipid metabolism,signal transduction and secondary metabolism,are affected.In order to clarify the mechanism of low temperature stress on the synthesis of phenolic substances in tobacco leaves and the response of tobacco seedlings to low temperature stress,the main results of this study are as follows:(1)Total polyphenol content was less affected by low temperature stress,and lignin content increased significantly(p<0.05);the activity of superoxide dismutase(SOD)was significantly increased(p<0.05),the activity of catalase(CAT)and total antioxidant capacity(T-AOC)decreased and lower than control treatment;PAL,HCT and POD enzyme activities was increased and the expression levels of PAL,HCT and POD gene were all increased under chilling stress.The above results indices that the metabolism of polyphenols shift to the downstream biosynthesis of lignin,and enhancing the hardness of cell walls by increasing lignin content is an important factor for plants to improve the plant's cold resistance.(2)Non-target method detected 642 kinds of GC/MS(gas chromatography/mass spectrometry)and stable metabolites,a total of 82 kinds of metabolic substances content is a marked change after low temperature treatment,mainly is the ramification of organic acids,amino acids,sugars,TCA cycle intermediate metabolites,etc.Under low temperature stress,the nitrogen metabolism of tobacco was enhanced,and the content of free amino acids and other nitrogen-containing substances in tobacco cells was increased.Under low temperature stress,the glucose metabolism pathway regulates cell infiltration and maintains cell water balance mainly by increasing soluble sugar content.Abscisic acid(ABA)content increased significantly under low temperature stress,which was beneficial to promote the signal transduction of plant under low temperature stress and to induce plants to respond to low temperature stress.(3)The up-regulated expressions of BAM,SUS and HxK in carbohydrate metabolism under low temperature conditions promoted the hydrolysis of starch,increased the content of soluble sugar in cells,and thus improved the cold resistance of plants.Plant hormone is the key factor of the regulation of plant response to low temperature stress,the ABA signal transduction genes PP2 C and rise in SnRK2 expression is involved in promoting the ABA signal transduction,to improve plant resistance at low temperature.During the amino acid metabolism,GST and GPx4 genes were up-regulated under low temperature stress,which enhanced the activity of phospholipid hydrogen peroxide glutathione peroxidase(GPx4)and glutathione s-transferase(GSTs),promoted the degradation of ROS,protected the integrity of membrane envelope system,and improved the low temperature resistance of tobacco.(4)Chilling stress increases the amount of polyphenols,especially lignin,which protects tobacco from chilling stress.An integrated analysis of metabolic and transcriptomic data revealed that most of the intermediate metabolites and enzymes related to lignin biosynthesis were synthesized in the leaves under chilling stress,which suggests that the biosynthesis of lignin plays an important role in the response of tobacco leaves to cold temperature.In addition,the cold insensitivity of CHS genes might be considered to be an important rate-limiting factor in the process of precursor substance flow to flavonoid biosynthesis under chilling stress.Furthermore,the up-regulated expression of PAL,HCT and CAD under chilling stress is the key to an increase in lignin synthesis.This study provides a hypothetical basis for the screening of new active metabolites and the metabolic engineering of polyphenols in tobacco.(5)Under the condition of low temperature stress,exogenous ABA treatment,such as SOD and CAT activity of antioxidant enzymes improve,improve the stability of the membrane system;ABA treatment significantly up-regulated the expression of key genes regulating lignin synthesis,such as PAL,HCT and CAD,and promoted the synthesis of lignin.The up-regulated expression of NtPYL8 promoted the binding of ABA to its receptor PYR/PYL/RCAR,thus inactivating PP2 C.SnRK2s was activated by its own phosphorylation and activated downstream targets of SnRK2 s phosphorylation,further amplifying the stress signal and enhancing the tolerance of plants to low temperature stress.