Metabolism of Sterols in Yeast

Parks, Leo W.; Adams, Bruce G.

doi:10.3109/10408417809090625

Cited by 184 publications

(83 citation statements)

References 255 publications

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“…The double bond at C-7, while not important for the sparking effect, is critical for polyene antibiotic sensitivity, which also depends on others factors, including phospholipid composition and presence of a suitable membrane potential (46). The particularly high level of resistance of the erg2 mutant (deficient in sterol ⌬8-⌬7 isomerase) confirms that the presence of ⌬5,7-conjugated double bond is determining.…”

Section: Discussionmentioning

confidence: 61%

Cloning by Metabolic Interference in Yeast and Enzymatic Characterization of Arabidopsis thaliana Sterol Δ7-Reductase

Lecain

Chenivesse

Spagnoli³

et al. 1996

Journal of Biological Chemistry

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Reduction of the ⌬7 double bond of sterols, a key biosynthetic step in higher eukaryotes, is lacking in lower eukaryotes like the yeast Saccharomyces cerevisiae, leading to terminal sterols with a ⌬5,7-conjugated diene structure. Genes encoding two sterol reductases involved, respectively, in the reduction of sterol ⌬14 and ⌬24(28) double bonds have been cloned to date, but no sequence information was available on the enzyme responsible for ⌬7-bond reduction. This study presents the cloning of the NADPH-sterol ⌬7-reductase (⌬7-red) from Arabidopsis thaliana, based on a metabolic interference approach in yeast. The principle is the functional expression of a plant cDNA library in the yeast strain FY1679-28C tolerant to sterol modifications and the selection of clones resistant to the polyene fungicide nystatin. The toxicity of this compound is dependent on the presence of ⌬5,7-unsaturated sterols in the yeast plasma membrane. One clone out of 10 5 transformants exhibits a cDNA-dependent alteration of cell sterol composition. The 1290-base pair cDNA open reading frame was isolated and sequenced. The corresponding protein presents a significant sequence similarity with yeast ⌬14-and ⌬24(28)-reductases and with human lamin B receptor. The coding sequence was extracted by polymerase chain reaction and inserted into a galactose-inducible yeast expression vector to optimize expression. Analysis using transformed wild type yeast or sterol altered mutants, indicated that ⌬5,7-ergosta-and cholesta-sterols are efficiently reduced in vivo, regardless of the structural variations on the side chain. No reductase activity was observed toward the ⌬14 or the ⌬5 positions of sterols. In vivo extensive ⌬7-reduction of the free and esterified pools of sterols was observed upon induction of the enzyme. Ergosterol present before induction was reduced into ergosta-5,22-dieneol, whereas ergosta-5-eneol is the new end product of sterol neosynthesis, indicating that the yeast ⌬22 desaturase may be no longer active on C-7-saturated sterols. In vitro tests indicated that ⌬7-reductase activity is preferentially associated with the endoplasmic reticulum membrane and confirmed the previous finding that NADPH is the reducing agent.

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Section: Discussionmentioning

confidence: 61%

Cloning by Metabolic Interference in Yeast and Enzymatic Characterization of Arabidopsis thaliana Sterol Δ7-Reductase

Lecain

Chenivesse

Spagnoli³

et al. 1996

Journal of Biological Chemistry

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“…This observation led to the view that Upc2p is activated in response to low heme levels (Abramova et al 2001b;Kwast et al 2002), as heme biosynthesis is tightly controlled by oxygen, and heme levels often serve as a surrogate for oxygen levels (Zhang and Hach 1999;Hon et al 2003). However, as ergosterol biosynthesis also requires oxygen (Parks 1978), hypoxic conditions would also lead to reduced sterol biosynthesis and might activate Upc2p by reducing sterol levels rather than heme levels.…”

Section: Resultsmentioning

confidence: 99%

A Role for Sterol Levels in Oxygen Sensing in Saccharomyces cerevisiae

Davies¹,

2006

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Upc2p and Ecm22p are a pair of transcription factors responsible for the basal and induced expression of genes encoding enzymes of ergosterol biosynthesis in yeast (ERG genes). Upc2p plays a second role as a regulator of hypoxically expressed genes. Both sterols and heme depend upon molecular oxygen for their synthesis, and thus the levels of both have the potential to act as indicators of the oxygen environment of cells. Hap1p is a heme-dependent transcription factor that both Upc2 and Ecm22p depend upon for basal level expression of ERG genes. However, induction of both ERG genes and the hypoxically expressed DAN/ TIR genes by Upc2p and Ecm22p occurred in response to sterol depletion rather than to heme depletion. Indeed, upon sterol depletion, Upc2p no longer required Hap1p to activate ERG genes. Mot3p, a broadly acting repressor/activator protein, was previously shown to repress ERG gene expression, but the mechanism was unclear. We established that Mot3p bound directly to Ecm22p and repressed Ecm22p-but not Upc2p-mediated gene induction.

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“…To summarize what is known about the metabolic or regulatory roles of sterols in several organisms, it has been found in one or more instances that specific sterols have some critical function in fatty acid transport and/or phospholipid biosynthesis (Holz et al, 1962;Dahl et al, 1980Dahl et al, , 1981Guyer & Bloch, 1983), and in respiration/mitochondrial activity (van Wagtendonk & Wulzen, 1950;Conner, 1957;Conner & Nakatani, 1958;Parks, 1978). For Paramecium, an apparent link between growth support and fatty acyl esterification of sterols has been reported (Conner et al, 1971).…”

Section: Discussionmentioning

confidence: 99%

Sterol Synergism in Paramecium tetraurelia

Whitaker

Nelson

1988

Microbiology

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Paramecium tetraurelia is a naturally occurring sterol auxotroph with an absolute nutritional requirement for one of a small group of structurally related phytosterols. We report here a quantitative study demonstrating that a low, otherwise sub-supportive, concentration ( x 0.020-0.050 pg ml-l) of an essential phytosterol (stigmasterol) is adequate for growth of this ciliate, provided that a second, relatively non-specific sterol is available at a higher concentration (1.0p.g ml-*) to allow for membrane biosynthesis. This phenomenon, referred to as sterol synergism, has been observed in a broad taxonomic range of organisms, with the conclusion that small amounts of specific sterols are required to perform some previously unknown, vital metabolic or regulatory function. Paramecium promises to be an excellent model organism for the elucidation of essential sterol function.

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Metabolism of Sterols in Yeast

Cited by 184 publications

References 255 publications

Cloning by Metabolic Interference in Yeast and Enzymatic Characterization of Arabidopsis thaliana Sterol Δ7-Reductase

Cloning by Metabolic Interference in Yeast and Enzymatic Characterization of Arabidopsis thaliana Sterol Δ7-Reductase

A Role for Sterol Levels in Oxygen Sensing in Saccharomyces cerevisiae

Sterol Synergism in Paramecium tetraurelia

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