Uptake and cellular distribution of copper, cobalt and cadmium in strains of Saccharomyces cerevisiae cultured on elevated concentrations of these metals

White, C

doi:10.1016/0378-1097(86)90003-0

Cited by 26 publications

(38 citation statements)

References 19 publications

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“…This distribution obtains up to 10 ~M -C U~+ in the medium. This result is somewhat different from the results of White & Gadd (1986) who recovered a larger fraction of intracellular copper in a particulate fraction (referred to as ' bound '). Intracellular uptake is energy-dependent, selective for Cu2+, negligible at 4 "C, and inhibited by cycloheximide (De Rome & Gadd, 1987;Lin & Kosman, 1990).…”

Section: Discussioncontrasting

confidence: 97%

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Distribution of 64Cu in Saccharomyces cerevisiae: cellular locale and metabolism

Lin

Crawford²,

Kosman³

1993

Journal of General Microbiology

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The metabolism of copper in the yeast Saccharomyces cerevisiae has been studied with respect to the distribution and stability to exchange of newly arrived @Cu. Cells pre-incubated with 10 ~M -C U~+ accumulated @Cu into two pools distinguishable by cellular locale and lability to exchange with extracellular cold copper. One pool was nonexchangeable and was localized to protoplasts. Size-exclusion chromatography of a soluble cell (protoplast) extract showed that this 64Cu was associated with up to four species. Two were identified as copper metallothionein and Cu,Zn superoxide dismutase based on comparisons of chromatograms derived from strains in which the genes for these two proteins had been deleted. A third species was identified as copper-glutathione based on chromatographic and biochemical assays. A second pool was exchangeable and was localized to the cell wall. In contrast to its rapid copper-stimulated exchange (t; M 1 min), this pool exhibited only slow efflux (10% 64Cu loss per 60 min). ZnZ+ did not stimulate the loss of 64Cu from this pool indicating that it was selective for copper, This pool was released into the supernatant upon protoplast formation and was found in the cell wall debris obtained when cells were mechanically disrupted. This 64Cu eluted in the void volume (peak P,) of the column used to sizefractionate copper-binding species. The metal in P, was exchangeable in vivo and in vitvo. However, the corresponding chromatographic fraction obtained from copper-naive cells when labelled in vitro could bind less than 20% of the @Cu bound to it in vivo indicating that the deposition of copper in this pool was primarily celldependent. In fact, this deposition was shown to be dependent on the cellular reduction of medium sulphate or sulphite to the level of sulphide, or on the addition of sulphide to the 64Cu uptake buffer. 64Cu in the nonexchangeable protoplast pool was not mobilized by cellular sulphide generation, indicating that cellular sulphide generation did not causally lead to the partitioning of @Cu to the cell wall pool. The data indicate that the appearance of copper sulphide(s) on the cell wall in S. cerevisiae is gratuitous and does not represent a sulphidebased mechanism of copper resistance in this yeast.

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Section: Discussioncontrasting

confidence: 97%

“…Some information about the overall distribution of metal has been published, however (White & Gadd, 1986). As noted, H,S generation has been proposed to causally lead to the deposition of copper in or on the cell wall, thus providing an important copper resistance mechanism in some yeasts (Kikuchi, 1965 ;Ashida, 1965).…”

Section: Introductionmentioning

confidence: 99%

Distribution of 64Cu in Saccharomyces cerevisiae: cellular locale and metabolism

Lin

Crawford²,

Kosman³

1993

Journal of General Microbiology

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“…The lack of copper efflux shows this strongly also (White & Gadd, 1986;Lin & Kosman, 1990;Lin et al, 1993). Thus, intracellular copper is not in thermodynamic equilibrium with respect to the medium.…”

Section: Discussionmentioning

confidence: 91%

“…1135-1141 and thus represents a fundamental aspect of normal yeast metabolism. Although the cellular response to elevated levels of copper and other transition metals has been studied in some detail in yeast and other micro-organisms (Kikuchi, 1965 ;Gadd &White, 1985;White & Gadd, 1986;Phelan et al, 1990;Greco et al, 1990), cellular copper handling by yeasts at normal copper levels has been characterized in somewhat less mechanistic detail (Wakatsuki et al, 1979(Wakatsuki et al, , 1988Gadd et al, 1984;De Rome & Gadd, 1987). The sulphide-dependent, periplasmic accumulation of copper could be easily dismissed as an artefact.…”

Section: Introductionmentioning

confidence: 99%

Distribution of 64Cu in Saccharomyces cerevisiae: kinetic analyses of partitioning

Lin

Crawford

Kosman

1993

Journal of General Microbiology

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The cell association of copper in the yeast Saccharomyces cerevisiae can involve both binding to the cell wall and the accumulation of copper within the cell. The former process requires the concurrent generation of H2S by the cell via the reduction of sulphate. The contributions of each of these processes to the uptake of V u by wild type and met3-containing (ATP sulphurylase-deficient) strains have been kinetically dissected. The Michaelis constant for uptake ( 4 p~) is independent of the type of cell association which is occurring, suggesting, although not requiring, that both processes are associated with a common kinetic intermediate. The time dependence of the cellassociation of 64Cu also suggests the presence of this intermediate pool of bound copper. The Vmax for uptake includes a constant contribution from accumulation of 64Cu within the plasmalemma [ O e l nmol min-' (mg protein)-'] plus that fraction of the 64Cu within the intermediate pool which diffuses away and is trapped on the cell wall as a metal sulphide. This latter contribution to V,,, can be two-to three-times greater than the intracellular uptake depending on the amount and type of sulphur supplementation provided in the 64Cu2+ uptake buffer. Both processes are energy-dependent although the sulphide-dependent periplasmic accumulation is somewhat more sensitive to metabolic inhibition. This can be attributed to the ATP required for the activation of sulphate prior to its reduction to the level of sulphite and then sulphide. Periplasmic 64Cu accumulation is strongly inhibited by Zn2+ and Ni2+. This inhibition is due to competition for cell-generated sulphide; in the presence of %n2+ , the decrease in 64Cu bound is quantitatively related to the amount of 65Zn which becomes cell-associated. In contrast, intracellular 64Cu uptake is not inhibited by these two metals (at 50 p~) showing that the copper translocation pathway is metalspecific. These observations suggest a model for the way newly arrived copper is handled at the cell membrane and is partitioned for intracellular uptake.

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“…In contrast to bacterial systems, in which toxic metals are pumped out of cells, toxic trace metals are often stored within yeast cells following transport across the plasma membrane (16). Excess metals that have gained access to the cytoplasm are sequestered by metal-binding proteins (40) or are compartmentalized within intracellular organelles such as the vacuole (45,61,62). The molecular biology of trace metal storage in yeast cells is best understood in the storage and detoxification of copper ions by the CUP1 gene product.…”

mentioning

confidence: 99%

COT1, a gene involved in cobalt accumulation in Saccharomyces cerevisiae.

Conklin¹,

McMaster²,

Culbertson³

et al. 1992

Mol. Cell. Biol.

189

126

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The COT] gene of Saccharomyces cerevisiae has been isolated as a dosage-dependent suppressor of cobalt toxicity. Overexpression (10,13,27). Although the CUP1 protein has been shown to bind several metals (64), increased expression of the CUP1 gene confers tolerance only to copper and cadmium (27, 30).We describe the isolation of COT1, a gene that when overexpressed enables wild-type strains of S. cerevisae to grow in media containing as much as 20 mM CoCl2. Deletion of the COTI gene makes cells more sensitive to cobalt than are wild-type strains. Since COT) also affects cobalt transport in a dosage-dependent manner, COT) appears to be involved in both the accumulation and detoxification of Co2+ ions. 3678on May 11, 2018 by guest

show abstract

Uptake and cellular distribution of copper, cobalt and cadmium in strains of Saccharomyces cerevisiae cultured on elevated concentrations of these metals

Cited by 26 publications

References 19 publications

Distribution of 64Cu in Saccharomyces cerevisiae: cellular locale and metabolism

Distribution of 64Cu in Saccharomyces cerevisiae: cellular locale and metabolism

Distribution of 64Cu in Saccharomyces cerevisiae: kinetic analyses of partitioning

COT1, a gene involved in cobalt accumulation in Saccharomyces cerevisiae.

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