Phytotoxic Effect Of Chromium Stress On Glandless Upland Cotton,its Alleviation And Comparative Study Of DNA Methylation And Transcriptome Profile

Glandless cotton can be grown to obtain cotton seeds free of toxic gossypol that will be used as food and feed for non-ruminant animals.Glandless cotton cultivars are not normally grown due to their susceptibility to various stresses.Chromium(Cr)is a potentially toxic heavy metal that has no basic metabolic function in plants,however,less information is available about the effects of Cr stress on glandless cotton.Despite the significance of Cr eco-toxicity,there is limited information and understanding gap about the mechanisms of Cr genotoxicity at the epigenetic level in plants as compared to humans and animals.Therefore,our study has also focused on DNA methylation and transcriptome changes and their relationship under Cr stress in different glanded and glandless cotton genotypes,to understand the molecular mechanism of cotton to Cr stress.In the present research,the four pairs of glanded and glandless cotton near-isogenic lines(NILs),ZMS17/ZMS-17w,ZMS-16/ZMS-16w,ZMS-12/ZMS-12w and Coker-312/Coker-312w were exposed to different doses(0,10,50,and 100?M/L)of Cr(VI)for seven days and examined their morphological,biochemical,ultrastructure,physiological,and molecular responses.From which it was observed that by increasing the level of Cr,inhibited seeds germination,and reduced the plant growth,fresh and dry biomass.The highest inhibition was shown in ZMS-16 and ZMS-17.Cr uptake in roots and translocation to stem and leaves were increased with the increasing Cr concentration.That was higher in roots and leaves mostly in glandless NILs while in glanded NILs it was higher in the stem.Impaired in nutrients uptake was also noted in different tissues of Cr treated plants.However,most of the nutrients uptake and translocation in glandless NILs were higher than their glanded NILs.Cr also caused ultrastructure damages in root tips,stem,and leaves showed large intercellular spaces,thick cell walls,distorted nucleus and chloroplast,ruptured nuclear membrane,large vacuole,damaged cell organelles,and electron-dense granules,which badly affects the biochemical,resulting reduction in the net photosynthesis rate,intercellular CO_2,and stomatal conductance.Thiobarbituric acid reactive substances(TBARS),H₂O₂increased with the increasing Cr concentration in all NILs and significantly activated the antioxidant enzymatic activities.Increasing the expression of genes encoding antioxidant enzymes both in Cr treated root and leaves contributes to the maintenance of the activity of antioxidant compounds.However,most of the genes expression of superoxide dismutase(SOD)and peroxidase(POD)showed similar trends with enzyme activities in glandless NILs.Differential responses of antioxidant enzymes and genetic levels to Cr stress indicated that antioxidant activities might play a predominant role in protection against Cr-induced oxidative stress.These results suggested that glanded and glandless cotton might perform different mechanisms to cope with Cr toxicity.The whole-genome DNA methylation in glanded and glandless cotton NILs and its correlation with gene expression by using whole-genome bisulfite sequencing(WGBS)under Cr stress were studied.Although,the relation of DNA methylation and gene expression under Cr stress in glanded and glandless cotton has poorly understood,the overall DNA methylation level within differentially methylated regions(DMRs)showed a significant increase under Cr stress,especially at the CHH context.However,the percentage of methylated cytosine at CG was higher and m CHH was the lowest.DMR associated gene was also higher at CHH context in the promotor region in all genotypes.Transcriptome analysis revealed that Cr suppressed the gene expression in cotton but the upregulated differentially expressed genes(DEGs)which were higher in ZMS-12 and TM-1 in response to Cr.Coker-312w not undergoes to high changes.Integration of DNA methylation and transcriptome analysis showed a positive correlation at CHH context in all genotypes,while negative at CHG.That indicated DNA methylation at CHG associated with gene repression and at CG had intermediate gene expression.Now it was clear from our results that DNA methylation at CHH context played an important role in gene upregulation under Cr stress in cotton.This study could provide new insight into understanding the molecular mechanisms of Cr tolerance in glanded and glandless cotton at the epigenetic level.This study also provides guidelines for future research in the breeding of Cr tolerant glandless cotton.Different abiotic stresses caused high phytotoxicity which disturbs the plants'normal growth and development.Therefore,it is necessary to use some alleviator which relieves the plants from abiotic stress caused phytotoxicity.An experiment was conducted to find out the alleviatory role of Sulphur(S),silicon(Si),and calcium(Ca)in glanded and glandless cotton NILs under Cr stress.The results showed that Cr stress-induced the morpho-physiological and biochemical changes in both glanded and glandless cotton NILs while their toxicity was alleviated by the application of alleviators(S,Si,and Ca).However,among the alleviators,S had better alleviatory results as compared to Si and Ca in all materials.Glandless cotton NILs showed high recovery from Cr toxicity in the application of alleviators with regards to all studied parameters as compared with their glanded NILs.The above results indicated that glandless cotton showed more tolerance to Cr than their glanded NILs and therefore was more appropriate for the Cr polluted environment.The morphophysiological ultrastructure and molecular responses could provide comprehensive insight into the response mechanism of glandless cotton induce by Cr stress.