The Role Of 5-aminolevulinic Acid On Regulation Mechanism Of Photosynthesis In Kentucky Bluegrass Seedlings Under Drought Stress

As one of the most common abiotic stress factors,drought causes negative effects on the physiological and metabolic processes of turfgrass species in arid and semi-arid lands.Following,the growth,aesthetic quality and seed yield of turfgrass can be considerably influenced by drought stress.Therefore,studies on the strategies and potential mechanisms of plant growth regulators?PGRs?to modulate the adverse effects of drought stress on turfgrass have become increasingly important for turfgrass establisment,maintenance and management.5-ALA?5-Aminolevulinic acid?is a potential PGR that is involved in regulating plant growth and yield under normal growth conditions and various abiotic stresses.However,the mechanisms of this regulation in response to stress,especially the molecular mechanisms,have not been fully documented.In addition,most of the reports on5-ALA-induced resistance were focused on crops,such as Brassica napus,Helianthus annuus,Triticum aestivum and Oryza sativa,the study on allexiating abiotic stress of turfgrass has been rarely reported.In the present study,the ptotective effects of exogenous 5-ALA on seed germination and seedlings growth were investigated in Kentucky bluegrass?Poa pratensis L.?,a widely used cool season turfgrass.Leaves of Kentucky bluegrass with 5-ALA pretreatment and drought stress were used for conducting transcriptome analyses based on RNA-Seq technology.The gene function and pathway of different expression genes were analysised by GO enrichment and KEGG pathway.Furthermore,physiological analyses on significantly enriched pathway were performed to demonstrate the results of transcriptional regulation.The following results were obtained?1?Seed soaking by 5-ALA at 0.1¹ mg·L^-1 had a promotional role on germination rate,germination potential,vigor inde,as well as plumule and radicle length under drought stress.The optimal concertration for seed soaking was 1 mg·L^-1.Foliar spray 5-ALA at 1²5 mg·L^-1 could alleviate drought-induced damage in Kentucky bluegrass seedlings.The leaf relative water content,turf quality and chlorophyll content were respectively increased by 41.9.2%,48.5%and 38.5%in plants with 5-ALA pretreatment at 10 mg·L^-1,while electrolyte leakage and MDA content were significantly less than plants with drought stress alone.?2?5-ALA pretreatment maintained ascorbate?AsA?and glutathione?GSH?contents and the ASA/DHA and GSH/GSSG ratios at high levels,and it enhanced the activities of superoxide dismutase?SOD?,catalase?CAT?,ascorbate peroxidase?APX?,glutathione peroxidase?GPX?,dehydroascorbate reductase?DHAR?,and glutathione reductase?GR?,which are crucial for scavenging drought-induced ROS.As a result,reactive oxygen species?ROS?production including H₂O₂ content and O₂^·-generation rate,lipoxygenase?LOX?activity,and malondialdehyde?MDA?content in plants with 5-ALA addition were reduced.?3?There were 3,021 differentially expressed genes?DEGs?following 5-ALA pretreatment?Drought+5-ALA?compared with the drought stress control?Drought alone?,including 1,317up-regulated and 1,704 down-regulated DEGs.To further characterize the function of DEGs,they were subjected to GO and KEGG enrichment analysis.According to GO and KEGG pathway enrichment analyses,photosynthesis,chlorophyll biosynthetic process,carbon fixation in photosynthetic organisms,response to water,glutamine biosynthetic process and carbohydrate metabolism were significantly enriched in biological processes and KEGG pathways.?4?Up-regulated expression of transcripts encoding Glutamyl-tRNA reductase?GluTR?and glutamate-1-semialdehyde 2,1-aminomutase?GSA-AT?were found in exogenous 5-ALA application plants,resulting in improveing 5-ALA biosynthesis and increasing endogenous 5-ALA content.These transcripts encode the key enzymes in chlorophyll biosynthesis progress from 5-ALA to chlorophyll a?Chl a?were also upregulated,including uroporphyrinogen decarboxylase,magnesium chelatase subunit H,magnesium-protoporphyrin IX monomethyl ester?oxidative?cyclase,and protochlorophyllide reductase.Additionally,5-ALA addition caused significant increases in the porphobilinogen?PBG?,-protoporphyrin IX?Proto IX?,magnesium-protoporphyrin IX?Mg-Proto IX?,protochlorophyllide?Pchl?and Chl a contents.?5?From KEGG pathway analysis,5-ALA application significantly regulated a series of proteins involved in photosystem II,including oxygen-evolving enhancer protein,10 kDa protein and 22 kDa protein,as well as Chlorophyll a/b-binding protein,then maintain the stability of light reaction center and increase PS II actual photochemical efficiency.Applied 5-ALA also resulted in the up-regulation of various transcripts encoding plastocyanin,ferredoxin and ferredoxin—NADP+reductase,which are related to photosynthethic electron transport.These results might be why 5-ALA maintained a higher electron transport rate?ETR?and photochemical quenching?qP?than did the drought stress control plants.Thermal dissipation and the over-reduction of QA could be reduced by higher ETR and qP,and lead to increasing net photothesis rate?Pn?.?6?5-ALA can cause an up-regulation of the expression of phosphoribulokinase and ribulose-bisphosphate carboxylase large chain under drought stress.In addition,the Rubisco activity in drought-stressed plants with 5-ALA pretreatment was significantly higher than that of plants without5-ALA pretreatment at 6 days after drought stress.This suggested that exogenous 5-ALA application could increase the phases of carbon fixation and ribulose-1,5-bisphosphate?RuBP?regeneration in the Calvin–Benson cycle.The expression of transcripts encoding triosephosphate isomerase?TPI?and fructose-1,6-bisphosphatase?FBP?was also up-regulated in response to 5-ALA treatment under drought stress.A significant increase in TPI and FBP activity in 5-ALA treatment plants with drought stress was consistently detected in this study.?7?The accumulation of sucrose and fructose involved in carbohydrate metabolism were inhabited by exogenous 5-ALA under drought stress.The increase of sucrose phosphatase?SPS?activity was also controlled by 5-ALA,while soluble acid invertase?SAI?and neutral invertase?NI?were improved.This results suggested that 5-ALA could decline the sucrose-induced feedback inhibition on photosynthesis in response to drought,as a result of regulating synthetase and catabolic enzyme activity to reduce the accumulation of sucrose in leaves of Kentucky bluegrass...