Construction of a set of convenient <i>saccharomyces cerevisiae</i> strains that are isogenic to S288C

Winston, Fred; Dollard, Catherine; Ricupero-Hovasse, S L

doi:10.1002/yea.320110107

Cited by 916 publications

(562 citation statements)

References 18 publications

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“…All JSY and ADM strains are derivatives of the FY10 strain (Winston et al, 1995). Standard genetic methods were used to grow, transform, and manipulate yeast (Sherman et al, 1986;Guthrie and Fink, 1991) and bacterial (Maniatis et al, 1982) strains.…”

Section: Yeast Strains and Methodsmentioning

confidence: 99%

The GTPase Effector Domain Sequence of the Dnm1p GTPase Regulates Self-Assembly and Controls a Rate-limiting Step in Mitochondrial Fission

Fukushima

Brisch

Keegan³

et al. 2001

MBoC

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Dnm1p belongs to a family of dynamin-related GTPases required to remodel different cellular membranes. In budding yeast, Dnm1p-containing complexes assemble on the cytoplasmic surface of the outer mitochondrial membrane at sites where mitochondrial tubules divide. Our previous genetic studies suggested that Dnm1p's GTPase activity was required for mitochondrial fission and that Dnm1p interacted with itself. In this study, we show that bacterially expressed Dnm1p can bind and hydrolyze GTP in vitro. Coimmunoprecipitation studies and yeast two-hybrid analysis suggest that Dnm1p oligomerizes in vivo. With the use of the yeast two-hybrid system, we show that this Dnm1p oligomerization is mediated, in part, by a C-terminal sequence related to the GTPase effector domain (GED) in dynamin. The Dnm1p interactions characterized here are similar to those reported for dynamin and dynamin-related proteins that form higher order structures in vivo, suggesting that Dnm1p assembles to form rings or collars that surround mitochondrial tubules. Based on previous findings, a K705A mutation in the Dnm1p GED is predicted to interfere with GTP hydrolysis, stabilize active Dnm1p-GTP, and stimulate a ratelimiting step in fission. Here we show that expression of the Dnm1 K705A protein in yeast enhances mitochondrial fission. Our results provide evidence that the GED region of a dynaminrelated protein modulates a rate-limiting step in membrane fission.

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Section: Yeast Strains and Methodsmentioning

confidence: 99%

The GTPase Effector Domain Sequence of the Dnm1p GTPase Regulates Self-Assembly and Controls a Rate-limiting Step in Mitochondrial Fission

Fukushima

Brisch

Keegan³

et al. 2001

MBoC

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“…Yeast strains are listed in Table 1 and are isogenic to S288C except that all strains are GAL2 ϩ (78). Lowercase indicates a mutant allele, and uppercase indicates a wild-type allele.…”

Section: Methodsmentioning

confidence: 99%

A New Class of Histone H2A Mutations in Saccharomyces cerevisiae Causes Specific Transcriptional Defects In Vivo

Hirschhorn

Bortvin

Ricupero-Hovasse

et al. 1995

Molecular and Cellular Biology

106

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Nucleosomes have been shown to repress transcription both in vitro and in vivo. However, the mechanisms by which this repression is overcome are only beginning to be understood. Recent evidence suggests that in the yeast Saccharomyces cerevisiae, many transcriptional activators require the SNF/SWI complex to overcome chromatin-mediated repression. We have identified a new class of mutations in the histone H2A-encoding gene HTA1 that causes transcriptional defects at the SNF/SWI-dependent gene SUC2. Some of the mutations are semidominant, and most of the predicted amino acid changes are in or near the N-and C-terminal regions of histone H2A. A deletion that removes the N-terminal tail of histone H2A also caused a decrease in SUC2 transcription. Strains carrying these histone mutations also exhibited defects in activation by LexA-GAL4, a SNF/SWI-dependent activator. However, these H2A mutants are phenotypically distinct from snf/swi mutants. First, not all SNF/SWI-dependent genes showed transcriptional defects in these histone mutants. Second, a suppressor of snf/swi mutations, spt6, did not suppress these histone mutations. Finally, unlike in snf/swi mutants, chromatin structure at the SUC2 promoter in these H2A mutants was in an active conformation. Thus, these H2A mutations seem to interfere with a transcription activation function downstream or independent of the SNF/SWI activity. Therefore, they may identify an additional step that is required to overcome repression by chromatin.In eukaryotic cells, DNA is complexed with histones and other proteins into chromatin (see reference 75 for a review). The primary component of chromatin is the nucleosome, which consists of approximately 146 bp of DNA wrapped around an octamer of histones (two histone H2A-H2B dimers and one [H3-H4] 2 tetramer; see reference 75 for a review). A growing body of evidence from both in vivo and in vitro studies has shown that the structure of chromatin influences gene expression (see references 25 and 53 for reviews). Biochemical experiments have shown that histones can repress transcription in vitro (see reference 53 for a review) and that transcriptional activators can overcome this repression (21,41,45,79,81,82). In vivo experiments in Saccharomyces cerevisiae have shown that the loss of transcriptional activators or of activator binding sites can be suppressed by mutations in histone genes (27,30,37,57). These results suggest that one function of transcriptional activators in vivo is to antagonize chromatin-mediated repression.In vivo studies of histone mutants have also suggested that histones play a variety of roles in transcriptional regulation. Small deletions and point mutations that alter the flexible N-terminal tails of different yeast histones have been shown to cause specific changes in transcription (see reference 25 for a review). Analysis of deletions and single amino acid changes in the N-terminal region of histone H4 has demonstrated that certain changes in the H4 N terminus abolish repression of the yeast silent mating-...

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“…23). The following strains were used in this study: JSY1781, MATa/MATα leu2Δ1/leu2Δ1 HIS3/his3Δ200 ura3-52/ura3-52DNM1-HA c /DNM1-HA c ; JSY3094, MATa leu2Δ1 his3Δ200 ura3-52 mdm20Δ::LEU2; JSY3095, MATa leu2Δ1 his3Δ200 ura3-52 mdm20Δ::LEU2 dnm1Δ::HIS3; JSY3096, MATα leu2Δ1 his3Δ200 ura3-52; JSY3097, MATα leu2Δ1 his3Δ200 ura3-52 dnm1Δ::HIS3; JSY3161, MATa his3Δ200 ura3-52 fzo1-1; JSY3162, MATα his3Δ200 ura3-52; JSY3163, MATa his3Δ200 ura3-52 fzo1-1 dnm1Δ::HIS3; and JSY3164 MATα his3Δ200 ura3-52 dnm1Δ::HIS3.…”

Section: Yeast Strainsmentioning

confidence: 99%

The dynamin-related GTPase Dnm1 regulates mitochondrial fission in yeast

et al. 1999

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The dynamin-related GTPase Dnm1 controls mitochondrial morphology in yeast. Here we show that dnm1 mutations convert the mitochondrial compartment into a planar 'net' of interconnected tubules. We propose that this net morphology results from a defect in mitochondrial fission. Immunogold labelling localizes Dnm1 to the cytoplasmic face of constricted mitochondrial tubules that appear to be dividing and to the ends of mitochondrial tubules that appear to have recently completed division. The activity of Dnm1 is epistatic to that of Fzo1, a GTPase in the outer mitochondrial membrane that regulates mitochondrial fusion. dnm1 mutations prevent mitochondrial fragmentation in fzo1 mutant strains. These findings indicate that Dnm1 regulates mitochondrial fission, assembling on the cytoplasmic face of mitochondrial tubules at sites at which division will occur.The mitochondrion is a complex organelle with a double membrane, its own genome and an independent protein-synthetic machinery. Although the role of mitochondria in metabolism and ATP production is more widely recognized, alterations in mitochondrial shape and abundance are also important for cellular function and differentiation. For example, mitochondrial morphology and copy number change in response to nutrient availability in yeast cells 1 , cell damage and apoptosis in mammalian cells 2 and developmental cues in Xenopus and Drosophila 3,4 . Cytological studies indicate that the 'steady-state' mitochondrial morphology and copy number can vary dramatically in different cell types, ranging from multiple, spherical organelles to the branched, tubular networks found in budding yeast and some mammalian cells [5][6][7] . These differences in mitochondrial morphology and copy number are largely determined by the balance between ongoing mitochondrial fission and fusion events.Little is known about molecules that regulate mitochondrial fission and fusion. The fuzzy onions (fzo) family of transmembrane GTPases was recently shown to control mitochondrial fusion in different organisms and cell types. In Drosophila, fzo is required for a developmentally regulated mitochondrial fusion event during spermatogenesis 8 . In budding yeast, mutations in fzo1 cause mitochondrial networks to fragment 9,10 and prevent mitochondrial fusion during yeast mating 9 . Molecules required for mitochondrial fission © 1999 Macmillan Magazines Ltd § Correspondence and requests for materials should be addressed to J.M.S. shaw@bioscience.utah.edu. NIH Public Access Results Mitochondrial membranes form nets in dnm1 mutant cellsWe previously reported that mitochondrial membranes collapse to one side of the cell in a dnm1Δ mutant strain 11 . Transmission electron microscopy indicated that these collapsed membranes might be organized in an unusual structure 11 . To characterize this structure further, we studied mitochondrial morphology in dnm1Δ cells under several different conditions.As reported previously, dnm1Δ mutants grown at 25 °C lack the highly branched mitochondrial network charac...

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Construction of a set of convenient saccharomyces cerevisiae strains that are isogenic to S288C

Cited by 916 publications

References 18 publications

The GTPase Effector Domain Sequence of the Dnm1p GTPase Regulates Self-Assembly and Controls a Rate-limiting Step in Mitochondrial Fission

The GTPase Effector Domain Sequence of the Dnm1p GTPase Regulates Self-Assembly and Controls a Rate-limiting Step in Mitochondrial Fission

A New Class of Histone H2A Mutations in Saccharomyces cerevisiae Causes Specific Transcriptional Defects In Vivo

The dynamin-related GTPase Dnm1 regulates mitochondrial fission in yeast

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