Study On The Mechanism Of Lipid Accumulation By Exogenous Oil In Mucor Circinelloides

Oleaginous microorganisms are capable of converting carbohydrates,hydrocarbons and common oils into their own intracellular lipids under specific conditions,and accumulate more than 20%lipids of their cell dry weight.Our group has found that Mucor circinelloides WJ11 is able to increase its lipid accumulation when using exogenous lipids as carbon source for fermentation,but the mechanism of lipid accumulation by exogenous lipids in M.circinelloides is not clear.In this thesis,the mechanism of lipid accumulation by M.circinelloides WJ11 using exogenous lipids,soybean oil,was investigated by transcriptomics analysis and metabolomics analysis.The main findings are as follows.(1)The optimal ratio of glucose-soybean oil mixed fermentation:without changing the carbon to nitrogen ratio of the original medium of M.circinelloides,the fermentation experiment was conducted using different ratios of soybean oil and glucose as a composite carbon source to replace the original glucose as the only carbon source.And the best mixture of glucose and soybean oil was selected based on the oil content accumulated by M.circinelloides.The fermentation results showed that the biomass and lipid accumulation of M.circinelloides the medium with the addition of exogenous soybean oil were higher than those of M.circinelloides grown in glucose as the carbon source.The optimal sugar-oil mixture ratio was 56.25%soybean oil and 43.75%glucose,i.e.35 g/L glucose and 24.3 g/L soybean oil,and the biomass of M.circinelloides under this fermentation condition was 14.99 g/L and the accumulated lipid content reached 43.83%,which was 17%higher than the control group.(2)The results of transcriptome analysis showed that a total of 4646 differentially expressed genes(DEGs)were identified in the treatment group(soybean oil and glucose mixture as carbon source)compared to the control group(glucose as a single carbon source)based on RNA-seq analysis.Among these genes,2379 genes were up-regulated and 2267genes were down-regulated in the treated group compared to the control group.The expression of genes of acetyl-Co A synthase(EC:6.2.1.1),6-phosphofructokinase(EC:2.7.1.11),alcohol dehydrogenase(NADP⁺),fructose bisphosphate aldolase,and pyruvate kinase involved in glycolysis changed significantly in the cells of M.circinelloides with soybean oil in the medium.The expression of two genes encoding acetyl-Co A synthase was down-regulated by 2^2.03-fold and 2^1.93-fold,respectively,and the expression of one gene encoding 6-phosphofructokinase was down-regulated by 2^1.74-fold.The expression of a gene encoding 6-phosphogluconate dehydrogenase(EC:1.1.1.49)was down-regulated by 2^1.78-fold.In contrast,many genes involved in lipid accumulation were up-regulated,including a 2^2.39-fold up-regulation of a gene,which encoding glycerol triphosphate O-acetyltransferase(3)Lipidomics analysis showed that a total of 438 different metabolites were identified.From the above-mentioned differential metabolites,81 significantly different metabolites were screened,of which 48 were up-regulated and 33 were down-regulated in M.circienelloides grown in mixed carbon source compared to that of M.circiellodes grown in glucose.These significantly different metabolites include phosphatidic acid(such as lyso-phosphatidic acid,phosphatidic acid,etc.),neutral glycerides(such as monoglycerides,diglycerides,etc.),fatty acids(such as(O-acyl)-1-hydroxy fatty acid),phosphatidylglycerol(such as lyso-phosphatidylglycerol,phosphatidylglycerol).Among them,the treatment group with the addition of soybean oil as a carbon source,lyso-phosphatidic acid and monoglyceride were up-regulated by 2⁵-fold and 2^4.48-fold,respectively,while phosphatidylglycerol and lyso-phosphatidylglycerol were down-regulated by 2^3.69-fold and 2^3.67-fold,respectively,compared to the control group with glucose only as a carbon source.