Mechanism Of Improving Chilling Adaptability Of Tomato Seedlings By Grafting Onto Cold-tolerant Rootstock

Low temperature is a common abiotic stress that severely affects growth and development of tomato,and ultimately reduces yield and fruit quality.Grafting is a technique that has been used since long to control biotic and abiotic stresses in different crop species.Some previous studies explained the physiological mechanism of rootstock to improve chilling tolerance of tomato,however,there is limited information available regarding molecular mechanism of rootstock to induce cold stress tolerance.In this study,two grafting combinations as LA4024/LA4024（self-grafted）and LA4024/LA1777（rootstock-grafted）were constructed using chilling tolerant‘LA1777’and chilling sensitive‘LA4024’tomato genotypes,and the mechanism of grafting to induce chilling tolerance of tomato was explored by physiological and transcriptomic analysis.The main results are as follows:1.Grafting onto chilling tolerant LA1777 rootstock can improve chilling adaptability of tomato seedlings.Grafting of chilling sensitive genotype onto chilling tolerant LA1777 rootstock restored the shoot turgor earlier and increased the relative water content at 4℃ temperature.This phenomenon might be attributed by the fact that the heterografted seedlings can maintain smaller stomatal aperture and stronger root activity under chilling stress.After 48 h of chilling treatment,the activities of SOD and POD of rootstock-grafted plants were stronger than those of self-grafted plants,and the accumulation of H₂O₂,O₂.^-and MDA was less,indicating less oxidative damage in rootstock-grafted plants caused by cold stress.2.Transcriptome sequencing was used to identify the differentially expressed genes（DEGs）of two grafting combinations.Transcriptomic sequencing of roots and leaves of grafted tomato plant was carried out after 0 h and 3 h of chilling treatment,and 3215 and 3795 DEGs were identified in the leaves of self-grafted and rootstock-grafted plants,respectively,at low temperature.Further KEGG enrichment analysis showed unique DEGs enrichment in more pathways in leaves of rootstock-grafted plants,indicating the rapid and diverse response of rootstock-grafted plants towards chilling stress.In addition,one Solyc09g008480.4 gene involved in phosphatidylinositol signaling system,two genes as Solyc10g076600.2 and Solyc04g054890.3 involved in jasmonic acid（JA）biosynthesis pathway,one Solyc12g013620.2 gene involved in JA signal pathway and regulation of stomatal aperture and some genes encoding transcription factors are found to be uniquely expressed in rootstock-grafted tomato plants.3.It was found that JA may be involved in the regulation of chilling tolerance of different grafted tomato combinations.The content of JA in self-grafted and rootstock-grafted plants were determined,and we observed significantly higher JA content in leaves of rootstock-grafted plants after 3-24 h of chilling treatment compared with self-grafted plants.The exogenous application of MeJA on self-grafted plants restored the shoot turgor and reduced the MDA accumulation.Whereas,the exogenous application of JA synthesis inhibitor SHAM on rootstock-grafted plants increased the MDA content.By analyzing the expression of key genes involved in JA synthesis and signalling pathway using q RT-PCR,it was observed that the expression of SlAOS,SlCOI1 and SlMYC2 genes in the leaves,and the expression of multiple JA synthesis pathway genes in the roots of rootstock-grafted plants was significantly higher than that of self-grafted plants after 3h chilling treatment.These results indicate that JA synthesis and signal pathways might be involved in the LA1777rootstock mediated improved chilling tolerance of tomato.Further,the expression of SlAOS gene was measured at multiple time points within 24 h of chilling treatment and the results verified the importance of this gene in inducing chilling tolerance.In addition,the expression level of SlCBF1/2 in the leaves of rootstock-grafted plants was higher than that of self-grafted plants under low temperature,indicating that higher endogenous JA accumulation in leaves of rootstock-grafted plants might induce cold tolerance by CBF pathway.