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Volumn 279, Issue 5353, 1998, Pages 1041-1044

Budding yeast Cdc20: A target of the spindle checkpoint

Author keywords

[No Author keywords available]

Indexed keywords

CELL CYCLE PROTEIN; CYCLINE;

EID: 0032512748     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.279.5353.1041     Document Type: Article
Times cited : (471)

References (35)
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    • (1995) Cell , vol.81 , pp. 279
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  • 18
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    • note
    • 1 arrest or with nocodazole (15 μg/ml) to activate the spindle checkpoint; in cdc26Δ strains, the cells were shifted to 37°C to inactivate the APC. Cells were harvested and lysed, and lysates were immunoprecipitated and protein immunoblotted as described (19), with the exception that the lysis buffer contained 50 mM β-glycerophosphate instead of 1 mM sodium vanadate.
  • 21
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    • unpublished data
    • A. Amon, unpublished data.
    • Amon, A.1
  • 24
    • 6844245402 scopus 로고    scopus 로고
    • note
    • The plasmid Ylplac211GAL-CDC20 contains the wild-type CDC20 open reading frame (ORF) fused to the GAL1 promoter and was directed to integrate into the URA3 gene by digestion with Nco I. To examine the spindle and DNA replication checkpoints, we grew wild-type cells (A1015), cells carrying two copies of a GAL-CDC20 fusion (A1016), or GAL-CDC20 cdc23-1 (A1022) cells to exponential phase and treated them with nocodazole (15 (μg/ml) or hydroxyurea (10 mg/ml) in YEP (yeast extract, peptone) raffinose at 23°C for 165 min to induce cell cycle arrest before adding galactose to 2% to induce production of excess CDC20. Samples were taken at the indicated times after galactose addition to determine DNA content and the percentage of cells that had rebudded. To examine the DNA damage checkpoint, we grew the cdc13-1 (A1017), cdc13-1 GAL-CDC20 (A1018), and cdc13-1 cdc23-1 GAL-CDC20 (A1023) strains to exponential phase at 23°C and shifted them to 33°C for 165 min in YEP raffinose medium before adding galactose. Samples were taken at the indicated times after galactose addition to determine DNA content and the fraction of cells that had rebudded. Viability was measured by adding nocodazole (15 μg/ml) or hydroxyurea (10 mg/ml), or shifting the temperature to 33°C (for cdc 13-1) for wild-type, mad1::URA3 (A928), mec1-1 (K2888) cdc13-1, or cdc13-1 rad9::URA3 (K2554) strains. For corresponding strains that also contained GAL-CDC20, galactose was added to 2% at the time of the other addition or temperature shift. Samples were withdrawn at the indicated times and plated on YEP glucose plates. Colonies were counted after 3 days. Induction of GAL-CDC20 leads to spindle elongation in cells arrested by the cdc13-1 mutation but not in cells arrested by hydroxyurea treatment (20,24).
  • 27
    • 6844248206 scopus 로고    scopus 로고
    • note
    • - glucose plates at the time of nocodazole addition and 2, 3.25, and 6 hours later.
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    • A.
    • Y. Yamamoto, A., V. Guacci, D. Koshland, J. Cell biol. 133, 99 (1996); O. Cohen-Fix and D. Koshland, Proc. Natl. Acad. Sci. U.S.A. 94, 14361 (1997).
    • (1996) J. Cell Biol. , vol.133 , pp. 99
    • Yamamoto, Y.1    Guacci, V.2    Koshland, D.3
  • 35
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    • note
    • We thank A. Rudner and A. Straight for yeast strains and plasmids, R.-H. Chen for antibodies to Mad2 and for disruption plasmids, A. Hoyt for communicating his unpublished results, and members of our labs for valuable discussions. We are grateful to T. Matsumoto for encouragement, helpful discussions, and for communicating unpublished results. Supported by a National Science Foundation Fellowship (L.H.H.), a grant from NIH (A.A.), and grants from NIH, the March of Dimes, and the David and Lucile Packard Foundation (A.W.M).


* 이 정보는 Elsevier사의 SCOPUS DB에서 KISTI가 분석하여 추출한 것입니다.