Effect Of Prohexadione-Calcium On Alleviating Saline-Alkali Stress In Soybean At Seedling Stage And Its Regulatory Mechanism

Soil salinization seriously restricts the growth and yield of soybeans.This study was conducted to investigate the mechanism of exogenous prohexadione-calcium(Pro-Ca)in alleviating saline-alkali stress of soybean at seedling stage.The salt-tolerant soybean variety’Hefeng 50’ and salt-sensitive soybean variety ’Kenfeng 16’ were used as experimental materials and the pot experiment was conducted to study the effects of foliar spraying 100 mg·L-1 Pro-Ca on morphogenesis,physiological characteristics,leaf ultrastructure,and transcriptome sequencing of soybean at seedling stage under 110 mmol L-1 saline-alkali stress.To reveal the physiological and molecular mechanism of Pro-Ca in alleviating saline-alkali stress.The main results and conclusions are as follows:1.With the extension of saline-alkali stress,the growth of the aerial part and roots of the two varieties of soybean seedlings were inhibited,and the difference was the most significant after 15 days of stress.Foliar spraying of Pro-Ca effectively alleviated the inhibition of saline-alkali stress on the growth parameters of two soybean seedlings and had a stronger regulation effect on saltsensitive variety Kenfeng 16,at day 15,Pro-Ca treatment significantly increased the root length and plant dry weight by 11.10%and 18.51%in Hefeng 50,respectively,and considerably increased the plant height,root length,plant fresh weight,and plant dry weight by 14.67%,19.65%,14.60%,and 14.05%in Kenfeng 16,respectively,compared to the saline-alkali treatment alone.2.Saline-alkali stress accelerated the decomposition of chlorophyll(Chl a,Chl b,Chl a+b,and Car)and significantly decreased the gas exchange parameters(Pn,Gs,Ci,and Tr)in the leaves of two soybean varieties.Moreover,saline-alkali treatment notably inhibited the increase of maximal photochemical efficiency of photosystem Ⅱ(Fv/Fm),potential activity of photosystem Ⅱ(Fv/Fo),and electron transport rate(ETR)in leaves of salt-sensitive variety Kenfeng 16.Foliar spraying of Pro-Ca significantly increased the contents of Chl a,Chl b,Chl a+b and Car,enhanced Fv/Fm,Fv/Fo,and ETR,maintained the activity of photosystem Ⅱ(PSⅡ),and in turn increased Pn,Gs,Ci and Tr in leaves of two soybean varieties under saline-alkali stress when compared to the saline-alkali treatment alone,and had a more significant effect on the salt-sensitive variety Kenfeng 16.3.Saline alkali treatment diminished the activities of some antioxidant enzymes in the late stage of stress,while suppressing the increase in AsA/DHA and GSH/GSSG ratios,resulting in a significant accumulation of ROS including superoxide anion((?))and hydrogen peroxide(H2O2)with treatment duration until the end of time course,which in turn led to an increase in malondialdehyde(MDA)and relative electrical conductivity(REC).Foliar spraying of Pro-Ca effectively scavenged ROS by further activated the activities of antioxidant enzymes(SOD,POD,CAT,PAL,and PPO),promoting the accumulation of osmotic solutes(soluble sugar,soluble protein,and proline),and increasing the ratio of AsA/DHA and GSH/GSSG,which in turn decreased MDA and REC levels.4.Under non-stress conditions,the leaf ultrastructure of two soybean varieties sprayed with or without Pro-Ca had a typical fusiform chloroplast shape,and the mitochondria were round and rich in the inner crest.Under saline-alkali stress,the chloroplast of the two soybean varieties were severely swollen,the grana lamellae were loosened,there were fewer starch grains and mitochondria and plenty of osmiophilic granules as compared to the control,and the degree of damage to chloroplasts was Kenfeng 16>Hefeng 50.Pro-Ca supplementation to the saline-alkali treated plants significantly alleviated the ultrastructural damage in leaves of two soybean varieties,repaired the thylakoid structure in chloroplast,increased the accumulation of starch grains,and inhibited the disintegration of mitochondria and the synthesis of osmiophilic granules.5.Transcriptome sequencing analysis showed that 2395(Hefeng 50)and 1618(Kenfeng 16)differentially expressed genes were identified under CKvs SA samples,and 770(Hefeng 50)and 938(Kenfeng 16)differentially expressed genes were detected under SA vs SA+Pro-Ca samples.KEGG analyses uncovered that the down-regulated differentially expressed genes in CK vs SA samples of the two varieties were mainly concentrated in phenylpropanoid biosynthesis,photosynthesis-antenna proteins,and flavonoid biosynthesis,indicating that the inhibition of photosynthesis and the decrease of ROS scavenging ability may be the main reasons for the inhibition of soybean seedling growth under saline-alkali stress.SA vs SA+Pro-Ca samples upregulated the co-expression of differentially expressed genes in ascorbate and aldarate metabolism and protein processing in endoplasmic reticulum,indicating that Pro-Ca can eliminate excessive ROS and stimulate growth by mediating the enhancement of antioxidant potential and protein modification.In conclusion,saline-alkali stress inhibited the growth of soybean seedlings and aggravated the damage of membrane lipid peroxidation.Exogenous Pro-Ca could enhance the saline-alkali tolerance of soybean at the seedling stage by delaying thylakoid membrane degradation,maintaining PSII activity,improving photosynthesis,enhancing antioxidant defense and osmotic regulation potential.The principal component analysis showed that Pro-Ca played an active role in responding to saline-alkali stress of different salt-tolerant soybean varieties.