Mechanisms Of Systemic Tolerance To Cold Stress Based On The Crosstalk Of Root And Shoot In Tomato Seedlings

Tomato(Solanum lycopersicum L.)is an important vegetable crop cultivated worldwide.Its growth,yield and quality can easily be affected by hostile environmental conditions like low temperature and weak light.At a whole plant level,there is a close correlation between the aboveground and belowground parts to the response towards the adverse conditions.Therefore,it’s of great significance to research how the low temperature systemic resistance can be regulated by the crosstalk between the aboveground and belowground parts.Grafting is a classic way to research signaling transduction.Using this approach,we can detect the involvement of signaling pathway between these parts while responding to low temperature stress.In this paper,we studied the resistant index,resistant gene expression and other aspects of low temperature treatment given to root/shoot induced systemic resistance in shoot/root of tomato plants along with possible involvement of ABA,RBOH1 via grafting.Therefore,present study was planned in three experiments to meet the desire objective.1.Effects of low temperature treatment to roots of tomato plants and the role of ABA in acquiring low temperature systemic resistance in shoots:Roots of tomato(Ailsa Craig;WT),were exposed to low temperature stress(4℃)by adding ice water to them.After giving the treatment,firstly,we examined the phenotype,relative electrical conductivity,stomatal movement,H2O2 content along with the differential expression of low temperature related resistant genes.We were also interested to know the role of ABA in tomato plants whose roots are exposed to low temperature stress.Hence,we studied the stomata development(density and sensitivity to ABA)in the leaves of WT and mutants(afterwards named as not)plants facing low temperature stress through roots.Thirdly,we also determined the gene’s expression levels in the leaves of grafted plants WT/not as well as in WT/WT in response to low temperature stress given to roots of tomato plants.It was observed from the study undertaken that the root-zone challenged with low temperature,lead to shoot wilting in a very short span of time,which was gradually restored afterwards.Low temperature stress also modulates the activity of stomata,which was observed to close first and then opened again.Relative conductivity and H2O2 content increased significantly with the onset of low temperature stress.The stomatal density in not plant was significantly higher than WT plant.Stomatal aperture was significantly increased,whereas stomatal sensitivity to ABA was decreased.It can be concluded that the stomatal movement could not be induced by root low temperature in not plant.Significant up-regulation in stress resistance related genes like ABA synthesis(NCED1,NCED4,NCED6),ABA signal regulating related gene(ABI5),CBF pathway related cold resistance gene(COR47),NADPH oxidase encoding gene(RBOH1),and HY5 signaling pathway genes(HY5 and COP1),and systemic resistance related gene(NPR1)was apparent facing to root-zone low temperature stress of WT/WT plants.There was no significant increase in above gene’s expression levels of WT/not plants at the same treatment.These results showed that the tomato roots facing low temperature treatment can induce resistance in the aerial part by regulating stomatal movement,expression of ABA,CBF,and HY5 pathway related genes,and accumulation of H2O2,which were possibly mediated by the root-derived ABA.2.Effects of low temperature to the shoots of tomato plants and the role of ABA and RBOH1 in acquiring low temperature systemic resistance in roots:The aboveground/shoot portions of tomato plants were exposed to 4℃ temperature in an artificial climate incubator,while controlling the root temperature at 25℃.After giving the required treatment,plants were screened for phenotype,stomatal movement and gene expression analysis for ABA biosynthesis pathway and CBF pathway.Secondly,we also carried the gene expression study for ABA synthesis and CBF-related gene expression in the roots of WT/WT,not/WT,rboh/WT grafted plants,WT as the rootstock,after exposing the aboveground portion to low temperature treatment for 12 hours.Thirdly,in order to understand the exact role of root-derived and leaf-derived ABA in tomato plants towards cold resistance,the grafted plants(WT/WT,WT/not,not/WT,not/not)were exposed to 4℃ low temperature for 7 days.Afterwards,phenotypes,relative conductivity,Fv/Fm and MDA content were studied.Results showed that the aboveground portion exposed to low temperature stress for the short period,did not change the relative conductivity of root.However,such duration of exposure is sufficient to induce stomatal closure,up-regulated root ABA synthesis genes(NCED1,NCED4,NCED6),ABA signal pathway regulation related genes(AREB,RD22-LIKE,SIT,AIM1 and FLA)and CBF pathway related genes(CBF1,CBF2,CBF3,COR47).The erasure of ABA and the silencing of RBOH1 gene in the aboveground part,significantly lower the expression ABA synthesis gene(NCED1,NCED6)and CBF-related genes(CBF1,CBF2 and CBF3)in the root;which was otherwise up-regulated in WT/WT,WT/not,not/WT,not/not grafted plants after 7 days of low temperature exposure.According to the phenotype,relative conductivity,Fv/Fm,MDA content,the resistances of the grafted plants decrease one by one.The above results showed that the effect of low temperature treatment to root is much lower than that of shoot within a short period of time,but sufficient enough to induce the expression of ABA pathway and CBF pathway related gene in roots.When the aboveground parts of plants are deficient in ABA or RBOH1 silenced,the induction in the expression of genes will either decreased or disappeared.It can be also concluded that both root-derived and leaf-derived ABA has played an important role in imparting cold resistance in tomato plants.3.Significance of light and ABA accumulation in roots of tomato plants induced by shoot low temperature treatment:Tomato plants(WT),were placed in the light/dark conditions along with shoot low temperature treatment.The expression of HY5,COP1,NCED1,NCED6 gene and the ABA content in the root of plants was studied after exposing them for 12 hours at 4/25℃.The results showed that the exposure of shoot at low temperature,significantly up-regulated expression of above genes in the light,but did not affect significantly under dark conditions.And significantly increased of root ABA content in the light,but not affect significantly under dark conditions.Hence,the induced expression of HY5 and ABA synthesis related genes and the ABA accumulation in roots might be correlated with light and were up-regulated in tomato plants when the shoot was exposed to low temperature.