Primary structure and transcription of an amplified genetic locus: the CUP1 locus of yeast.

Karin, Michael; Najarian, Rick; Haslinger, Alois; Valenzuela, Pablo; Welch, Juliet W.; Fogel, Seymour

doi:10.1073/pnas.81.2.337

Cited by 269 publications

(167 citation statements)

References 39 publications

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“…The first two C residues in the CUP1 transcript were converted to G by site-directed mutagenesis and a HpaI site was added at ϩ25 (relative to the first major transcript start site; ref. 23) to give pCUP1C-less (30). A synthetic 380-bp C-less cassette was obtained as a blunted AseI-SacI fragment from pB20A inserted at HpaI͞SacI to yield pCUP1C-less(410).…”

Section: Methodsmentioning

confidence: 99%

An initiation element in the yeastCUP1promoter is recognized by RNA polymerase II in the absence of TATA box-binding protein if the DNA is negatively supercoiled

LeBlanc

Benham

Clark

2000

Proc. Natl. Acad. Sci. U.S.A.

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Purified RNA polymerase II initiated transcription from the yeast CUP1 promoter fused to a C-less cassette if the DNA was negatively supercoiled. Relaxed plasmid was not transcribed. Transcription did not require addition of any other transcription factors. TATA box-binding protein (TBP) was not detectable in the polymerase preparation and the TATA box was not required. Deletion analysis of the CUP1 promoter revealed that a 25-bp element containing the initiation region was sufficient for recognition by polymerase. Two transcription start sites were mapped, one of which is identical to one of the two major start sites observed in vivo. Our observations can be accounted for by using a theoretical analysis of the probability of DNA melting within the plasmid as a function of superhelix density: the CUP1 initiation element is intrinsically unstable to superhelical stress, permitting entry of the polymerase, which then scans the DNA to locate the start site. In support of this analysis, the CUP1 promoter was sensitive to mung bean nuclease. These observations and a previous theoretical analysis of yeast genes support the idea that promoters are stress points within the DNA superhelix. The role of transcription factors might be to mark the promoter and to regulate specific melting of promoter DNA.

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Section: Methodsmentioning

confidence: 99%

An initiation element in the yeastCUP1promoter is recognized by RNA polymerase II in the absence of TATA box-binding protein if the DNA is negatively supercoiled

LeBlanc

Benham

Clark

2000

Proc. Natl. Acad. Sci. U.S.A.

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“…cerevisiae contains a single Cu-MT gene which is present in the CUPl locus located on chromosome VIII (Fogel & Welch, 1982;Karin et al, 1984). Copper-resistant S. cerevisiae strains (CUPl'') contain 2-14 or more copies of the CUPl locus, which are randomly repeated, and may grow on media containing ^ 2 mM copper (Fogel, Welch & Karin, 1983;Karin et al, 1984). Copper-sensitive strains (cupP) do not grow at concentrations ^0-15mM (Butt & Ecker, 1987).…”

Section: Intracellular Fate Of Toxic Metals* (A) Metal-binding Proteimentioning

confidence: 99%

Interactions of fungi with toxic metals

1993

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SUMMARY This review details the major interactions of fungi with toxic metal species. Physico‐chemical and biological aspects are discussed including definitions and classification of metals in biological contexts, mechanisms of metal fungitoxicity, resistance and tolerance, environmental influences and the occurrence of fungi in metal‐polluted habitats. Cellular interactions include extracellular precipitation and complexation, binding to cell walls, transport of monovalent and divalent metal cations, intracellular fates of toxic metal species (including metal‐binding proteins and peptides, and vacuolar compartmentation), and metal transformations including organometal(loid) synthesis. Significant interactions between metals and mycorrhizal fungi and macrofungi are summarized with reference to the amelioration of plant phytotoxicity and metal accumulation respectively. Biotechnological aspects of metal‐fungal interactions relate to the biological treatment of metal‐contaminated effluents and waste streams for metal removal/recovery and environmental protection.

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“…CcO levels vary depending on whether the cells are grown by fermentation or respiration. In addition, a varying quantity of cellular copper exists bound to two metallothioneins, Cup1 and Crs5 (7,8). Expression of CUP1 and CRS5 is regulated by copper levels through the copper-responsive transcription factor Ace1 (9,10).…”

mentioning

confidence: 99%

Yeast Contain a Non-proteinaceous Pool of Copper in the Mitochondrial Matrix

Cobine

Ojeda

Rigby

et al. 2004

Journal of Biological Chemistry

207

199

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The yeast mitochondrion is shown to contain a pool of copper that is distinct from that associated with the two known mitochondrial cuproenzymes, superoxide dismutase (Sod1) and cytochrome c oxidase (CcO) and the copper-binding CcO assembly proteins Cox11, Cox17, and Sco1. Only a small fraction of mitochondrial copper is associated with these cuproproteins. The bulk of the remainder is localized within the matrix as a soluble, anionic, low molecular weight complex. The identity of the matrix copper ligand is unknown, but the bulk of the matrix copper fraction is not protein-bound. The mitochondrial copper pool is dynamic, responding to changes in the cytosolic copper level. The addition of copper salts to the growth medium leads to an increase in mitochondrial copper, yet the expansion of this matrix pool does not induce any respiration defects. The matrix copper pool is accessible to a heterologous cuproenzyme. Co-localization of human Sod1 and the metallochaperone CCS within the mitochondrial matrix results in suppression of growth defects of sod2⌬ cells. However, in the absence of CCS within the matrix, the activation of human Sod1 can be achieved by the addition of copper salts to the growth medium.Copper is an essential cell nutrient acting as a cofactor in nearly 20 enzymes (1). However, excess accumulation of copper ions results in toxicity. Evidence for the effectiveness of copper ions as a toxin comes from its historic use as a fungicide, molluscide, and algicide. Homeostatic mechanisms exist in cells to regulate the cellular concentration of copper ions, thus maintaining copper balance and minimizing deleterious effects. Cells appear to maintain a quota for essential metal ions; this quota is primarily the quantity necessary to metallate the various copper proteins (2, 3). Copper ions are required for at least three key enzymes in Saccharomyces cerevisiae. The cuproenzymes include the cytosolic superoxide dismutase Sod1, 1 the plasma membrane ferroxidase Fet3, and the mitochondrial inner membrane enzyme cytochrome c oxidase (CcO). The copper quota of the yeast S. cerevisiae is about 5 ϫ 10 5 atoms per cell (3, 4).It is unclear what fraction of the 5 ϫ 10 5 copper atoms per cell is from copper in Sod1, Fet3, and CcO. Expression of these copper-binding proteins varies with growth conditions, suggesting that the copper may be distributed differently depending on growth conditions. Clearly, a significant fraction of the cellular copper is associated with Sod1; however, not all Sod1 molecules are metallated (4, 5). Fet3 requires four copper ions for activity, but levels of this protein are dependent on iron status of the medium (6). CcO levels vary depending on whether the cells are grown by fermentation or respiration. In addition, a varying quantity of cellular copper exists bound to two metallothioneins, Cup1 and Crs5 (7,8). Expression of CUP1 and CRS5 is regulated by copper levels through the copper-responsive transcription factor Ace1 (9, 10). An increase in the free Cu(I) ion pool activates Ace1 throu...

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Primary structure and transcription of an amplified genetic locus: the CUP1 locus of yeast.

Abstract: Copper resistance in yeast is controlled by the CUP) locus. The level of resistance is proportional to the

Cited by 269 publications

References 39 publications

An initiation element in the yeastCUP1promoter is recognized by RNA polymerase II in the absence of TATA box-binding protein if the DNA is negatively supercoiled

An initiation element in the yeastCUP1promoter is recognized by RNA polymerase II in the absence of TATA box-binding protein if the DNA is negatively supercoiled

Interactions of fungi with toxic metals

Yeast Contain a Non-proteinaceous Pool of Copper in the Mitochondrial Matrix

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