Role Of GSNOR In Disease Resistance To Botrytis Cinerea Induced By GABA In Tomato

Tomato,rich in vitamins,lycopene and other nutritional and functional ingredients,is popular among consumers.However,tomato fruit is easily infected by Botrytis cinerea after harvest and causesing a lot of losses.Therefore,it is of great significance to explore a green,safe,broad-spectrum,efficient method for reducing postharvest diseases in fruits and vegetables.Gamma-aminobutyric acid(GABA)is a four-carbon amino acid,widely distributes in the free state of prokaryotes and eukaryotes,and plays an important role in mammals and plants.However,there are few studies on its role in postharvest disease control of fruits and vegetables.S-nitroso glutathione reductase(GSNOR),a member of alcohol deoxidizing enzyme family in plants,can regulate nitric oxide(NO)and its metabolites nitroso mercaptans(SNOs)levels as a key enzyme.It was involved in plant growth,against pathogenic bacteria,heat tolerance,cell death cadmium stress and a variety of physiological processes.In this study,we used a GSNOR inhibitor-N6022,transient silencing and stable inheritance methods to explore the role of GSNOR in GABA-induced resistance to Botrytis cinerea in tomato fruit.The specific results are as follows:(1)The incidence and disease area of tomato fruit were significantly(P < 0.05)reduced by 10 m M GABA solution.Moreover,the activity of GSNOR and gene expression in fruits were significantly(P < 0.05)increased,and the level of NO was significantly(P < 0.05)increased within 12 h after treatment,but it was significantly lower(P < 0.05)than that of the control group after inoculation for 8 h with Botrytis cinerea.(2)In tomato fruit treated with 60 ?M GSNOR inhibitor N6022,the development of disease spots was accelerated,the disease resistance was weakened,and the content of NO maintained in a higher level compared with the control,and the enzyme activity and gene expression of GSNOR were significantly(P < 0.05)inhibited.(3)After GSNOR of tomato was transient silenced by virus-induced gene silencing(VIGS)technique and then inoculated by B.cinerea,the endogenous NO and SNOs was significant(P < 0.05)accumulated,the enzyme activity and gene expression of GSNOR were significantly(P < 0.05)reduced,and the disease resistance of tomato fruit became weaker.After the exogenous addition of GABA,the levels of NO and SNOs were alleviated,and the enzyme activity and gene expression level of GSNOR were significantly(P < 0.05)activated,as well as the disease resistance was enhanced.(4)Agrobacterium-mediated tomato transformation technology was used for instantaneous overexpressing or silencing GSNOR.Ninteen overexpression positive plants were obtained,of which the highest expression level of GSNOR was about 13 times that of wild type(WT).After expansion and cultivation,it was found that GSNOR enzyme activity and GSNOR gene expression were significantly(P < 0.05)inhibited in RNAi tomato,resulting in excessive accumulation of NO and SNOs and weaker disease resistance to Botrytis cinerea.The content of NO and SNOs in plants overexpressing Sl GSNOR was lower than that of the wild type,indicating that overexpression of Sl GSNOR can metabolize excess NO and SNOs in the plant,avoid damage to the cell,and induce the pathogenesis-related protein 1 gene(PR1)and non-expressor gene(NPR1),the marker genes of systemic acquired resistance in plant.The results suggest that overexpression of Sl GSNOR is benefit for improving the disease resistance of tomato to Botrytis cinerea.(5)Subcellular localization showed that Sl GSNOR protein was localized on the intima of guard cells,indicating that Sl GSNOR protein play an important role in improving the defensive ability of tomato plant.