Genome-Wide Analysis of Sterol-Lipid Storage and Trafficking in Saccharomyces cerevisiae

ABSTRACT The pandemic of lipid-related disease necessitates a determination of how cholesterol and other lipids are transported and stored within cells. The first step in this determination is the identification of the genes involved in these transport and storage processes. Using genome-wide screens, we identified 56 yeast (Saccharomyces cerevisiae) genes involved in sterol-lipid biosynthesis, intracellular trafficking, and/or neutral-lipid storage. Direct biochemical and cytological examination of mutant cells revealed an unanticipated link between secretory protein glycosylation and triacylglycerol (TAG)/steryl ester (SE) synthesis for the storage of lipids. Together with the analysis of other deletion mutants, these results suggested at least two distinct events for the biogenesis of lipid storage particles: a step affecting neutral-lipid synthesis, generating the lipid core of storage particles, and another step for particle assembly. In addition to the lipid storage mutants, we identified mutations that affect the localization of unesterified sterols, which are normally concentrated in the plasma membrane. These findings implicated phospholipase C and the protein phosphatase Ptc1p in the regulation of sterol distribution within cells. This study identified novel sterol-related genes that define several distinct processes maintaining sterol homeostasis.

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Analysis of Neutral Lipid Synthesis in Saccharomyces cerevisiae by Metabolic Labeling and Thin Layer Chromatography

Journal of Visualized Experiments ◽

10.3791/62201 ◽

2021 ◽

Author(s):

Sean Rogers ◽

W. Mike Henne

Keyword(s):

Saccharomyces Cerevisiae ◽

Thin Layer ◽

Neutral Lipid ◽

Thin Layer Chromatography ◽

Lipid Synthesis ◽

Metabolic Labeling ◽

Layer Chromatography

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Identification of novel genes involved in neutral lipid storage by quantitative trait loci analysis of Saccharomyces cerevisiae

BMC Genomics ◽

10.1186/s12864-021-07417-4 ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Klavdija Pačnik ◽

Mojca Ogrizović ◽

Matthias Diepold ◽

Tobias Eisenberg ◽

Mia Žganjar ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Lipid Metabolism ◽

Quantitative Trait Loci ◽

Neutral Lipid ◽

Lipid Content ◽

Quantitative Trait ◽

Lipid Accumulation ◽

Lipid Storage ◽

Neutral Lipid Content ◽

Trait Loci

Abstract Background The accumulation of intracellular fat depots is a polygenic trait. Therefore, the extent of lipid storage in the individuals of a species covers a broad range and is determined by many genetic factors. Quantitative trait loci analysis can be used to identify those genetic differences between two strains of the same species that are responsible for the differences in a given phenotype. We used this method and complementary approaches to identify genes in the yeast Saccharomyces cerevisiae that are involved in neutral lipid storage. Results We selected two yeast strains, the laboratory strain BY4741 and the wine yeast AWRI1631, with a more than two-fold difference in neutral lipid content. After crossing, sporulation and germination, we used fluorescence activated cell sorting to isolate a subpopulation of cells with the highest neutral lipid content from the pool of segregants. Whole genome sequencing of this subpopulation and of the unsorted pool of segregants implicated several loci that are involved in lipid accumulation. Three of the identified genes, PIG1, PHO23 and RML2, were investigated in more detail. Deletions of these genes and the exchange of the alleles between the two parental strains confirmed that the encoded proteins contribute to neutral lipid storage in S. cerevisiae and that PIG1, PHO23 and RML2 are the major causative genes. Backcrossing of one of the segregants with the parental strains for seven generations revealed additional regions in the genomes of both strains with potential causative genes for the high lipid accumulation phenotype. Conclusions We identified several genes that contribute to the phenotype of lipid accumulation in an allele-specific manner. Surprisingly, no allelic variations of genes with known functions in lipid metabolism were found, indicating that the level of storage lipid accumulation is determined by many cellular processes that are not directly related to lipid metabolism.

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