Regulatory Mechanisms Of Exogenous Phytohormone On Lipid Accumulation In Foodborne Coccomyxa Subellipsoidea

Plant hormone is an important small molecular signal substance that regulates cell growth and metabolism,and plays the key role in regulating the biomass accumulation and intracellular lipid accumulation of microalgae.However,the specific molecular mechanism of its induction of intracellular lipid synthesis and accumulation in microalgae is not clear.In this paper,Coccomyxa subellipsoidea C-169 was cultured by salicylic acid,methyl jasmonate and abscisic acid.The effects of three exogenous plant hormones on the growth and lipid accumulation of Coccomyxa subellipsoidea were studied,and the regulation mechanism of lipid accumulation induced by the best exogenous hormones was preliminarily analyzed.The main results are as follows:1)Different concentrations of salicylic acid(SA),methyl jasmonate(Me JA)and abscisic acid(ABA)were added to the culture system of Coccomyxa subellipsoidea at the initial growth stage(0 h)and stable stage(168 h),respectively.The results showed that the highest lipid yield was 3.05 g/L when cultured for 216 h with 20 mg/L SA at the initial growth stage,which was 19.73% higher than that of the control group.The highest lipid yield obtained by adding 5 mg/L Me JA for 48 hours in the growth stable period was 2.89 g/L,which was 13.21% higher than that in the control group,and the highest lipid yield obtained by adding 8 mg/L ABA in the growth stable period for 48 hours was 3.16 g/L,which was 24.02% higher than that in the control group.Therefore,when 8 mg/L ABA was added in the stable growth period,the lipid yield increased the most after 48 hours of induction.2)Based on the optimal lipid accumulation induction conditions of Coccomyxa subellipsoidea,the cells treated with 8 mg/L ABA for 48 h were used as the experimental samples,and the algae cells without ABA as the control samples for transcriptome analysis.The results showed that among the 1102 differentially expressed genes(DEGs),568 were significantly down-regulated and 534 were significantly upregulated.The expression pattern of q RT-PCR,with Gene 39868,42677,54573 and83467 was the same as that of the transcriptional group.It is confirmed that the transcriptional group is more reliable to screen the data of differentially expressed genes.Under the induction of exogenous ABA,up-regulation of DEGs was mainly concentrated in lipid metabolism and amino acid metabolism pathway,while downregulation of DEGs was mainly located in energy metabolism,cofactor and vitamin metabolism pathway.38 key DEGs were selected to regulate lipid synthesis and accumulation in Coccomyxa subellipsoidea.Based on GO and KEGG annotation and enrichment analysis,38 key differential genes in plant hormone signal transduction pathway were further screened,among which ABA signal pathway genes(Gene 64904,25912 and 47867),auxin signal pathway genes(Gene 59830,40946,65085 and 59751),ethylene pathway gene 52299 and ROS signal pathway genes(Gene 37867 and 15220)were significantly differentially expressed.3)The prediction of promoter homeopathic elements and expression correlation analysis of 38 key genes(KEYGENEs)in upstream of hormone signal pathway and 38 key regulatory DEGs in downstream lipid synthesis pathway of Coccomyxa subellipsoidea confirmed that the upstream genes of plant hormone signal pathway induced by exogenous ABA can affect the expression pattern of key genes in downstream lipid accumulation pathway,thus regulating lipid accumulation in Coccomyxa subellipsoidea.Then,based on the upstream KEYGENEs of hormone signal pathway and downstream lipid synthesis pathway DEGs,the intracellular lipid accumulation pathway of Coccomyxa subellipsoidea induced by exogenous ABA was constructed,and the activities of regulatory enzymes PGK,GPD,ME,FAS,GDPD,MGAT and DGAT related to the pathway of differentially expressed genes regulating lipid accumulation were characterized.It was found that the PGK activity of algae cells induced by ABA was 33.46% lower than that of the control samples without ABA induction.The activities of GPD,ME,FAS,GDPD,MGAT and DGAT were upregulated by 14.75%,11.07%,23.88%,17.68%,42.95% and 47.82% respectively compared with the control,which was consistent with the expression pattern of these pathway-related regulatory genes.It was confirmed that under the induction of exogenous ABA,the photosynthetic carbon fixation pathway regulated by these enzyme genes was weakened and the fatty acid pathway was strengthened,which made the carbohydrate metabolic pathway flow to the lipid pathway.In addition,the enhanced glyceride pathway provides energy and precursors for the synthesis and accumulation of TAG,thus promoting the intracellular lipid accumulation of Coccomyxa subellipsoidea.