PAH1-Mediated Regulation of the Transcriptome and Lipidome in Yarrowia lipolytica
The PAH1 gene encodes for phosphatidate phosphatase (PAP), which plays a vital role in lipid homeostasis. PAP catalyzes the conversion of phosphatidate (PA) to diacylglycerol (DAG) used to synthesize triacylglycerol (TAG). Research with Saccharomyces cerevisiae outlined the Pah1 fundamental role in...
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| Format: | Dissertation |
| Language: | English |
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ProQuest Dissertations & Theses
01-01-2022
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| Online Access: | Get full text |
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| Summary: | The PAH1 gene encodes for phosphatidate phosphatase (PAP), which plays a vital role in lipid homeostasis. PAP catalyzes the conversion of phosphatidate (PA) to diacylglycerol (DAG) used to synthesize triacylglycerol (TAG). Research with Saccharomyces cerevisiae outlined the Pah1 fundamental role in lipid metabolism. Pah1 in S. cerevisiae serves many functions, such as lipid droplet formation, vacuole homeostasis, and its role in transcriptional regulation of genes involved in phospholipid synthesis. However, there have been limited studies with Pah1 in the oleaginous yeast Yarrowia lipolytica. Conclusions about Y. lipolytica cannot be made solely on research with S. cerevisiae because the steps for lipid biosynthesis are not the same. The objective of this research is to elucidate the role of PAH1 in Y. lipolytica grown in media that favors lipid accumulation. Yarrowia lipolytica wild-type (WT) and pah1Δ cells that lack PAH1 were grown in high glucose. The PAP activity was measured following the release of water-soluble 32Pi from chloroform-soluble [32P] PA. Lipids were analyzed by TLC, GC, and LC/MS. The expression of the Pah1 was analyzed by immunoblot analysis and by qPCR. RNAseq was used for global gene expression analysis. The results showed a significant difference in the growth and biomass production; lipid analysis showed the pah1Δ still synthesized TAGs but lower than the WT strain. The reduced TAG amounts in the pah1Δ were balanced by increased phospholipids and free fatty acids (FFA). The PAP activity increased in a time-dependent; however, this did not correlate with an increase in Pah1 levels. The qPCR results showed a decrease in Pah1 transcript correlating with the reduction in protein levels in the membrane fraction. The transcriptome analysis revealed several differentially expressed genes between the WT and pah1Δ. This study also looked at the effect of the nem1Δ on Y. lipolytica cells. There were no significant differences in the WT and nem1Δ until day 4, which outlined asubstantial decrease in DAGs and an increase in TAG. Overall, this study sought to elucidate the role of PAH1 in Y. lipolytica. These results can help further understand the functional role of PAH1 in the lipid biosynthetic pathway. |
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| ISBN: | 9798379920708 |