Yeast mutants auxotrophic for choline or ethanolamine

Atkinson, K D; Jensen, Birgitte Nybo; Kolat, A I; Storm, Eva; Henry, Susan A.; Fogel, Seymour

doi:10.1128/jb.141.2.558-564.1980

Cited by 174 publications

(67 citation statements)

References 37 publications

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“…Therefore, it is not surprising that some auxotrophic mutations and/or their required nutritional supplements affect sporulation efficiency (e.g. see Atkinson et al, 1980;Freese et al, 1984;Schroeder and Breitenbach, 1981;Varma et al, 1985). In addition, auxotrophy interferes with formation of pseudohyphae; that is, pseudohyphal formation is best observed with prototrophic strains in minimal medium with no added nutritional supplements (J. Heitman, personal communication).…”

Section: Discussionmentioning

confidence: 99%

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Three new dominant drug resistance cassettes for gene disruption in Saccharomyces cerevisiae

Goldstein¹,

McCusker²

1999

Yeast

654

838

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Disruption-deletion cassettes are powerful tools used to study gene function in many organisms, including Saccharomyces cerevisiae. Perhaps the most widely useful of these are the heterologous dominant drug resistance cassettes, which use antibiotic resistance genes from bacteria and fungi as selectable markers. We have created three new dominant drug resistance cassettes by replacing the kanamycin resistance (kan(r)) open reading frame from the kanMX3 and kanMX4 disruption-deletion cassettes (Wach et al., 1994) with open reading frames conferring resistance to the antibiotics hygromycin B (hph), nourseothricin (nat) and bialaphos (pat). The new cassettes, pAG25 (natMX4), pAG29 (patMX4), pAG31 (patMX3), pAG32 (hphMX4), pAG34 (hphMX3) and pAG35 (natMX3), are cloned into pFA6, and so are in all other respects identical to pFA6-kanMX3 and pFA6-kanMX4. Most tools and techniques used with the kanMX plasmids can also be used with the hph, nat and patMX containing plasmids. These new heterologous dominant drug resistance cassettes have unique antibiotic resistance phenotypes and do not affect growth when inserted into the ho locus. These attributes make the cassettes ideally suited for creating S. cerevisiae strains with multiple mutations within a single strain.

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Section: Discussionmentioning

confidence: 99%

“…However, the extensive use of auxotrophic mutations is unfortunate because it limits analysis to auxotrophic strains and because auxotrophies are frequently deleterious, (e.g. see Atkinson et al, 1980;Baganz et al, 1997;Schroeder and Breitenbach, 1981).…”

Section: Introductionmentioning

confidence: 99%

Three new dominant drug resistance cassettes for gene disruption in Saccharomyces cerevisiae

Goldstein¹,

McCusker²

1999

Yeast

654

838

View full text Add to dashboard Cite

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“…Cultures were grown at 30ЊC in YPGE in the presence of 32 P i (50 Ci ml ¹1 ) to steady-state labelling as described previously (Atkinson et al, 1980). Spheroplasts were prepared as follows.…”

Section: Phospholipid Analysismentioning

confidence: 99%

Cardiolipin is not essential for the growth of Saccharomyces cerevisiae on fermentable or non‐fermentable carbon sources

Jiang

Rizavi

Greenberg

1997

Molecular Microbiology

173

143

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SummaryCardiolipin is a unique dimeric phospholipid, which is present throughout the eukaryotic kingdom and is specifically localized in mitochondrial membranes. It is widely believed that mitochondria possess an essential requirement for this phospholipid. To determine whether cardiolipin is essential for yeast growth, we generated a cardiolipin synthase null mutant by disrupting the CLS1 gene (open reading frame YDL142c on chromosome IV) of Saccharomyces cerevisiae. Biochemical analysis of the mutant indicated that it had no cardiolipin synthase activity and no cardiolipin in its membranes. The enzyme phosphatidylglycerolphosphate synthase, which catalyses the committed step of the cardiolipin pathway, remained unaffected in the null mutant. Haploid cells containing the null allele are viable in media containing glucose, galactose or glycerol/ethanol as the sole carbon source, although growth in galactose or glycerol/ethanol is somewhat reduced in the mutant compared with the wild type. These results indicate that cardiolipin is not essential for the growth of S. cerevisiae in fermentable or nonfermentable carbon sources.

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“…Cells were labelled overnight, for at least six generations, with 5 lCiAEmL )1 of 32 P-labelled orthophosphate in 5 mL of vitamin-defined synthetic I + Cmedia, and harvested during the late logarithmic phase of growth. Labelled lipids were extracted as described by Atkinson et al [29]. 2D paper chromatography on silicaimpregnated paper was performed according to the method of Steiner & Lester [30].…”

Section: Phospholipid Compositionmentioning

confidence: 99%

Regulation of phospholipid biosynthesis by phosphatidylinositol transfer protein Sec14p and its homologues

Holic̆

Zagorsek

Griač

2004

European Journal of Biochemistry

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Transcription of yeast phospholipid biosynthesis structural genes, which contain an inositol-sensitive upstream activating sequence in their promoters, responds to the availability of the soluble precursors inositol and choline and to changes in phospholipid metabolism. The INO1 gene is deregulated (derepressed when inositol is present) under the conditions of increased phosphatidylcholine (PtdCho) turnover, as occurs in the sec14D cki1D strain (SEC14 encodes the major yeast phosphatidylinositol transfer protein; CKI1 encodes choline kinase of the cytidine diphosphate choline pathway of PtdCho biosynthesis). Five proteins (Sfhp) share sequence homology with phosphatidylinositol transfer protein Sec14p. Two (Sfh2p and Sfh4p), when overexpressed largely complement the otherwise essential Sec14p requirement concerning growth and secretion. In this study, we analysed the ability of Sec14 homologues to correct the defect in regulation of phospholipid biosynthesis resulting from defective or missing Sec14p. We also analysed how PtdCho turnover relates to the transcriptional regulation of phospholipid biosynthesis. The results show that (a) none of the Sec14 homologues was able to substitute for Sec14p in its regulatory aspects of phospholipid biosynthesis, (b) removal of phospholipase D activity corrected the aberrant INO1 gene regulation in yeast strains with otherwise high PtdCho turnover, and (c) increased steady-state phosphatidic acid levels correlated with derepressed levels of the INO1 gene. Overall, the results support the model in which high phosphatidic acid levels lead to derepression of the genes of phospholipid biosynthesis [Henry, S

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Yeast mutants auxotrophic for choline or ethanolamine

Cited by 174 publications

References 37 publications

Three new dominant drug resistance cassettes for gene disruption in Saccharomyces cerevisiae

Three new dominant drug resistance cassettes for gene disruption in Saccharomyces cerevisiae

Cardiolipin is not essential for the growth of Saccharomyces cerevisiae on fermentable or non‐fermentable carbon sources

Regulation of phospholipid biosynthesis by phosphatidylinositol transfer protein Sec14p and its homologues

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