Abstract
[Objective]The aim of this study was to investigate the effects of exogenous N-carbonyloc-tanoyl homoserine lactone(3OC8-HSL)on salt tolerance of Malus robusta seedlings by evaluating growth performance,physiological parameters,and the expression of salt-responsive genes under salt stress.[Methods]A greenhouse pot experiment was conducted with four treatment groups:Including the control group(CK,0 mmol·L-1 NaCl+water),the 3OC8-HSL treatment group(AC,0 mmol·L-1 NaCl+10 μmol·L-1 3OC8-HSL),salt stress group(Salt,200 mmol·L-1 NaCl+water),and 3OC8-HSL+salt stress group(AS,200 mmol·L-1 NaCl+10 μmol·L-1 3OC8-HSL).Each treatment was applied to the seedlings for 45 days.3OC8-HSL solution(10 μmol·L-1)was drench-applied to the root zone on days 1,7,and 14,while salt stress was imposed by watering with 200 mmol·L-1 NaCl every 3 days.The growth parameters(plant height and fresh mass)were measured at the end of the experiment.The leaf chlorophyll content was determined(total chlorophyll,as well as chlorophyll a and b)as an indicator for photosynthetic status.The malondialdehyde(MDA)content in seedling roots was quantified to as-sess lipid peroxidation(oxidative stress damage).The leaf Na+and K+concentrations were measured,and the Na+/K+ratio was calculated to evaluate ionic homeostasis under each treatment.For molecular analysis,the root samples from salt-stressed seedlings with and without 3OC8-HSL were subjected to RNA sequencing(RNA-seq).Differentially expressed genes(DEGs)between the AS treatment and Salt treatment were identified using DESeq2(standard:|log2 fold change|＞1 and adjusted p-value＜0.05).The gene Ontology(GO)enrichment analysis was performed to characterize the biological processes en-riched among these DEGs.Furthermore,four DEGs associated with stress responses were selected for validation by quantitative real-time PCR(RT-qPCR):MdPAL,MdGST,MdHKT1,and MdLRR-RLK.[Results]The Growth and Physiological Responses:Under non-saline conditions,the exogenous 3OC8-HSL markedly promoted the growth of Malus robusta seedlings.The treated seedlings exhibited a 23.74%increase in plant height and a 17.23%increase in fresh mass compared with the untreated con-trols,along with a slight but significant rise in the leaf chlorophyll content(+4.06%).These results in-dicated that 3OC8-HSL itself had a growth-promoting effect even without stress.Under salt stress(200 mmol·L-1 NaCl),3OC8-HSL application substantially improved seedling performance and stress tolerance.The 3OC8-HSL-treated salt-stressed seedlings grew taller(+11.61%plant height)and heavier(+26.38%fresh mass than those under salt stress alone.Notably,3OC8-HSL alleviated salt-induced damage:the treated seedlings maintained significantly higher chlorophyll levels(total chlorophyll+36.98%vs salt control),indicating better photosynthetic capacity under stress.In addition,3OC8-HSL reduced oxidative damage,as evidenced by a 36.20%reduction in root MDA accumulation under salt stress relative to untreated seedlings.This lower MDA level suggested that 3OC8-HSL would enhance the antioxidant defense,thereby protecting cellular membranes from peroxidation damage caused by salt stress.The Ion Homeostasis:3OC8-HSL also improved ionic balance in Malus robusta under salini-ty.In the salt-stressed seedlings,3OC8-HSL treatment led to significantly lower Na+accumulation and higher K+retention in plant tissues.Specifically,the Na+content in the 3OC8-HSL-treated seedlings was 11.01%lower than that in the untreated salt-stressed controls,while the K+content was 19.46%higher.Consequently,The Na+/K+ratio,a key indicator of ion homeostasis under salt stress,was re-duced by 22.17%in the 3OC8-HSL group compared with the salt treatment alone.This improvement in Na+/K+homeostasis reflected a better maintenance of ionic equilibrium,which is crucial for salt toler-ance.The results suggested that 3OC8-HSL would help seedlings limit sodium uptake or transport and/or enhance potassium uptake,thereby mitigating ion toxicity under high salinity.Consistently,the treat-ed seedlings showed healthier morphology and less salt injury than the untreated ones(greener leaves,less wilting),as observed qualitatively during the experiment.The Gene Expression and Pathway Acti-vation:The transcriptome profiling of roots under salt stress revealed significant molecular changes due to3OC8-HSL.A total of 560 genes were differentially expressed in the 3OC8-HSL-treated vs.untreat-ed salt-stressed roots,with 177 genes upregulated and 383 downregulated(adjusted P＜0.05).The GO enrichment analysis of these DEGs indicated that 3OC8-HSL activated multiple stress-responsive path-ways.Many of the upregulated genes were associated with secondary metabolite biosynthesis and de-fense-related processes.In particular,biological process categories such as phenylpropanoid biosynthet-ic and metabolic pathways(involved in the synthesis of lignin and other phenolics),glutathione metabo-lism,response to wounding,and cuticle development were significantly enriched among the 3OC8-HSL-induced genes.These results implied that 3OC8-HSL would trigger a broad reprogramming of the plant's stress response at the molecular level,enhancing both structural and chemical defenses under the salt stress.Importantly,several key salt-tolerance genes were strongly upregulated by 3OC8-HSL treatment.For instance,transcripts of the MdPAL(phenylalanine ammonia-lyase,a rate-limiting enzyme in the phenylpropanoid pathway leading to lignin synthesis)increased 4.6-fold in the 3OC8-HSL-treat-ed roots(after 7 days of salt stress)compared with the untreated controls.The MdHKT1,encoding a high-affinity K+transporter involved in Na+/K+homeostasis,was upregulated 9.6-fold,and the MdLRR-RLK,encoding a leucine-rich repeat receptor-like kinase implicated in stress signal perception,rose by 8.4-fold(both after 7 days).Additionally,the MdGST(glutathione S-transferase,involved in detoxifica-tion and ROS scavenging via the glutathione pathway)showed a 6.4-fold induction at 24 hours after treatment.The robust induction of these genes,which would play roles in lignin biosynthesis(cell wall fortification),antioxidant activity,ionic transport,and stress signaling,would underscore the multi-fac-eted mechanism by which 3OC8-HSL would enhance salt tolerance.[Conclusion]This study demon-strated that the exogenous 3OC8-HSL improved the salt tolerance of Malus robusta seedlings by pro-moting growth,enhancing photosynthetic capacity,maintaining Na+/K+homeostasis,and reducing oxi-dative damage.The transcriptome and RT-qPCR analysis showed that the 3OC8-HSL upregulated the expression of the MdPAL,MdGST,MdHKT1,and MdLRR-RLK,and activated pathways related to phen-ylpropanoid metabolism and glutathione metabolism under the salt stress.These results would provide a theoretical basis for the application of AHL signals in improving salt resistance of apple rootstocks.关键词
八棱海棠/盐胁迫/N-羰基辛酰基高丝氨酸内酯/Na+/K+平衡Key words
Malus robusta/Salt stress/N-octanoyl-L-homoserine lactone/Na+/K+homeostasis分类
农业科技
