Transcriptome Analysis Combined With Lipid Metabolome Analysis Reveals The Triacylglycerol Synthesis Mechanism In Cyperus Esculentus

Cyperus esculentus is a perennial herb native to tropical and subtropical regions of Africa,Asia,and Europe.It widely distribute throughout the world.Cyperus esculentus tubers are rich in lipids and have extremely exploitable potential in the vegetable oil industry.In this study,transcriptomic analysis and lipid metabolomic analysis were performed to investigate the lipids accumulation rules of the tubers of Cyperus esculentus at four different developmental stages(TD-1,TD-2,TD-3,and TD-4),and to establish a molecular pathway for the triacylglycerol synthesis of Cyperus esculentus.The specific findings are as follows:1.Transcriptome sequencing of Cyperus esculentus tubers yielded a total of181356 genes at four different developmental stages by a next-generation sequencing platform.Of these,121865 were functionally annotated.The numbers of differentially expressed genes(DEGs)between different developmental stages were as follows:There were 4013 between TD-1 and TD-2,of which 1984 were up-regulated and 2029 were down regulated;There were 1295 DEGs between TD-2 and TD-3,of which 1984 were up-regulated and 2029 were down regulated;There were 14349 DEGs between TD-3 and TD-4,of which 1984 were up-regulated and 2029 were down regulated.This result indicated that there were more differed expression genes t at the later developmental stage in the tubers of Cyperus esculentus compared to the pre developmental stage.The differential expression genes related to lipids synthesis were enriched in different developmental periods of Cyperus esculentus tubers.Eight and ten lipids metabolic pathways were enriched by GO term and KEGG term respectively.Total 86 transcription factor families which had different expression level under different developmental period were identified based on transcriptome analysis.Of these,45 transcription factor families were identified in TD-1 versus TD-2;26transcription factor families were identified in TD-2 versus TD-3;70 transcription factor families were identified in TD-3 versus TD-4.The expression of transcription factors,MYB96-1、MYB96-2、MYB41-1、MYB41-2、DOF-1、DOF-2、DOF-3 和DOF-4 were gradually downregulated,and the expression of transcription factors,DOF-5、DOF-6、WRI1-3、WRI1-4、WRI1-5、WRI1-6、WRI1-7、WRI1-8、WRI1-9、WRI1-10、WRI1-11 were gradually upregulated with the development of tubers.The expressing rules of these genes have consistence with the accumulation rules of FA or TAG.2.255 lipid classes were identified by lipid metabolome analysis in the different developmental stages tubers of Cyperus esculentus,including 144 glycerolipids(7subclasses),71 glycerophospholipids(9 subclasses),sphingolipids(5 subclasses),and10 fatty acids.It was found that the glycerolipid content continuously was accumulated according to tuber development,and increased faster at later developmental stages than that at earlier stages.Triacylglycerol content accounted for over 71.5% of the total glyceride in the ripening tubers.Accumulation trend of triacylglycerol was consistent with the total glyceride content.The content of glycerophospholipids increased first and then decreased according to the tuber development.Phosphatidic acid(PA)accounted for the highest percentage,25.7% of glycerophospholipids in the ripening tubers.The total sphingomyelin content was consistently low at different developmental periods;Fatty acids continuously decreased according to tuber development,which may be related to the rapid accumulation of TAG content at later stages of development.To explore changes in fat metabolism dependent on tuber developmental stage,we performed a differential metabolite(DM)analysis.The results showed that a total of188 differential metabolites belonged to glycerolipid and glycerophospholipid metabolic pathways mostly,were identified in adjacent developmental stages.Among them,97 differential metabolites were identified between TD-1 and TD-2(1 was upregulated and 97 downregulated);TD-2 VS TD-3 had 82 differential metabolites(38was upregulated and 44 downregulated);TD-3 VS TD-4 had 146 differential metabolites(37 was upregulated and 109 downregulated).3.Analysis combined transcriptome and lipid metabolome showed that critical genes for FA synthesis,ACCase-1、ACCase-2 、ACCase-3、MCAT、KASⅠ-1、KASⅠ-2、KASⅠ-3、KASⅠ-4、ENR-1、ENR-2、ENR-3、KASⅡ-1、KASⅡ-2、KASⅡ-3、FATB-1、FATB-2、FATB-3、SAD-3、FATA-2、FATA-3,and the key genes involved in TAG synthesis,GPAT9、LPAAT1-4、DGAT3-1、DGAT3-2,were up-regulated depending on tuber development,which is consistent with the pattern of oil accumulation.The oleosin genes and the caleosin genes are involved in the formation of oil bodies.In this study,we identified 9 oleosin genes and 21 caleosin genes,which remained high level expression,except CeCLE-8 and CeCLE-20.