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Volumn 285, Issue 5427, 1999, Pages 591-595

Genetic selection of peptide inhibitors of biological pathways

Author keywords

[No Author keywords available]

Indexed keywords

CARRIER PROTEIN; NUCLEASE;

EID: 0033597939     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.285.5427.591     Document Type: Article
Times cited : (174)

References (43)
  • 1
    • 0021818675 scopus 로고
    • G. P. Smith, Science 228, 1315 (1985); S. Cwirla, E. A. Peters, R. W. Barrett, W. J. Dower, Proc. Natl. Acad. Sci. U.S.A. 87, 6378 (1990); J. K. Scott and G. P. Smith, Science 249, 386 (1990); J. J. Devlin, L. C. Panganiban, P. E. Devlin, ibid., p. 404.
    • (1985) Science , vol.228 , pp. 1315
    • Smith, G.P.1
  • 3
    • 0025112794 scopus 로고
    • G. P. Smith, Science 228, 1315 (1985); S. Cwirla, E. A. Peters, R. W. Barrett, W. J. Dower, Proc. Natl. Acad. Sci. U.S.A. 87, 6378 (1990); J. K. Scott and G. P. Smith, Science 249, 386 (1990); J. J. Devlin, L. C. Panganiban, P. E. Devlin, ibid., p. 404.
    • (1990) Science , vol.249 , pp. 386
    • Scott, J.K.1    Smith, G.P.2
  • 4
    • 0025004285 scopus 로고    scopus 로고
    • G. P. Smith, Science 228, 1315 (1985); S. Cwirla, E. A. Peters, R. W. Barrett, W. J. Dower, Proc. Natl. Acad. Sci. U.S.A. 87, 6378 (1990); J. K. Scott and G. P. Smith, Science 249, 386 (1990); J. J. Devlin, L. C. Panganiban, P. E. Devlin, ibid., p. 404.
    • Science , pp. 404
    • Devlin, J.J.1    Panganiban, L.C.2    Devlin, P.E.3
  • 5
    • 0029876415 scopus 로고    scopus 로고
    • P. Colas et al., Nature 380, 548 (1996).
    • (1996) Nature , vol.380 , pp. 548
    • Colas, P.1
  • 10
    • 0344796158 scopus 로고    scopus 로고
    • note
    • 16GGTGGTGTCGACAC-3′ [N = A:T:G:C (1:1:1:1), S = G:C (1:1)]. Annealed DNA was extended by using the klenow fragment of DNA polymerase I, and the double-stranded product was cut with Eco RI and Sal I to create two equivalents of library DNA that were ligated to the staphylococcal nuclease construct (pTCN22) cut with Eco RI and Sal I, installing library DNA in-frame.
  • 11
    • 0024285092 scopus 로고
    • The expression vector pTCN23 carried the replicator of the 2-μm circle, an endogenous yeast high-copy plasmid, the PGK1 promoter [ A. Chambers, C. Stanway, A. J. Kingsman, S. M. Kingsman, Nucleic Acids Res. 16, 8245 (1988)], and had codon usage optimized according to Sharp and Li [P. M. Sharp and W. H. Li, Nucleic Acids Res. 15, 1281 (1987).
    • (1988) Nucleic Acids Res. , vol.16 , pp. 8245
    • Chambers, A.1    Stanway, C.2    Kingsman, A.J.3    Kingsman, S.M.4
  • 12
    • 0023650543 scopus 로고
    • The expression vector pTCN23 carried the replicator of the 2-μm circle, an endogenous yeast high-copy plasmid, the PGK1 promoter [ A. Chambers, C. Stanway, A. J. Kingsman, S. M. Kingsman, Nucleic Acids Res. 16, 8245 (1988)], and had codon usage optimized according to Sharp and Li [P. M. Sharp and W. H. Li, Nucleic Acids Res. 15, 1281 (1987).
    • (1987) Nucleic Acids Res. , vol.15 , pp. 1281
    • Sharp, P.M.1    Li, W.H.2
  • 15
    • 0028872743 scopus 로고
    • X. Li and R. B. Nicklas, Nature 373, 630 (1995); C. L. Rieder, R. W. Cole, A. Khodjakov, G. Sluder, J. Cell Biol. 130, 941 (1995); W. A. Wells and A. W. Murray, ibid. 133, 75 (1996).
    • (1995) Nature , vol.373 , pp. 630
    • Li, X.1    Nicklas, R.B.2
  • 17
    • 0029912971 scopus 로고    scopus 로고
    • X. Li and R. B. Nicklas, Nature 373, 630 (1995); C. L. Rieder, R. W. Cole, A. Khodjakov, G. Sluder, J. Cell Biol. 130, 941 (1995); W. A. Wells and A. W. Murray, ibid. 133, 75 (1996).
    • (1996) J. Cell Biol. , vol.133 , pp. 75
    • Wells, W.A.1    Murray, A.W.2
  • 18
    • 0032546360 scopus 로고    scopus 로고
    • D. P. Cahill, et al., Nature 392, 300 (1998).
    • (1998) Nature , vol.392 , pp. 300
    • Cahill, D.P.1
  • 20
    • 0025941405 scopus 로고
    • M. A. Hoyt, L. Totis, B. T. Roberts, Cell 66, 507 (1991); R. Li and A. W. Murray, ibid., p. 519 (1991).
    • (1991) Cell , pp. 519
    • Li, R.1    Murray, A.W.2
  • 22
    • 0345227047 scopus 로고    scopus 로고
    • note
    • Unless otherwise indicated, all experiments in this study were done in strains isogenic to W303. A single selection strain, DA2050A (a/a bar1/bar1 mec1-1/ mec1-1 CDC28VF::LEU2/CDC28-VF::LEU2 GAL1-MPS1/CAL1-MPS1), was engineered to identify inhibitors of the spindle checkpoint or of the pheromone response pathway. The bar1 mutation increases sensitivity to α factor by removing a protease that degrades the pheromone, and the CDC28-VF mutation blocks the ability of cells to adapt to overexpression of Mps1 from the GAL1 promoter. The mec1-1 mutation, which abolishes the DNA damage checkpoint, is irrelevant to the experiments reported here.
  • 23
    • 0345658913 scopus 로고    scopus 로고
    • note
    • For both selections, transformants were grown on uracil-deficient plates for 2 days. Cells were scraped from these plates, and aliquots were plated onto selection plates [lacking uracil and containing galactose (-uracil/+galactose) for the spindle checkpoint selection; lacking uracil and containing α factor (-uracil/+α factor) (1 μg/ml) for the pheromone pathway selection] and grown at 30°C for 2 or 3 days. Plasmid DNA was recovered from the entire population of colonies that grew on these plates, purified, amplified by transformation into E. coli, and transformed into DA2050A to enrich for peptamers that conferred resistance as opposed to genomic mutations that caused resistance. Cell populations harboring active peptamers typically grew as lawns on selection plates, and individual plasmids were isolated from these plates and retested to confirm that they gave plasmid-dependent resistance.
  • 28
    • 0031658303 scopus 로고    scopus 로고
    • M. S. Singer et al., Genetics 150, 613 (1998).
    • (1998) Genetics , vol.150 , pp. 613
    • Singer, M.S.1
  • 38
    • 0345227039 scopus 로고    scopus 로고
    • note
    • Abbreviations for the amino acid residues are as follows: A, Ala; C, Cys; D, Asp; E, Glu; F, Phe; G, Gly; H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; R, Arg; S, Ser; T, Thr; V, Val; W, Trp; and Y, Tyr.
  • 39
    • 0030455820 scopus 로고    scopus 로고
    • A yeast ORF-Gal4 activation domain fusion array has been assembled that expresses about 85 to 90% of the predicted ORFs of 5. cerevisiae in strain pJ69-4a (P. James, J. Halladay, E. Craig Genetics 144, 1425 (1996)]. In this strain, GAL4 is absent and the reporters HIS3 and ADE2 are under Gal4 control. To probe the Gal4-AD fusion array for protein-protein interactions, we mated the array to a strain expressing a Gal-4-DNA binding domain fusion. After selecting for diploids, we identified two-hybrid positives by testing diploids on plates containing different concentrations of 3-aminotriazole.
    • (1996) Genetics , vol.144 , pp. 1425
    • James, P.1    Halladay, J.2    Craig, E.3
  • 40
    • 0032512748 scopus 로고    scopus 로고
    • L. H. Hwang et al., Science 279, 1041 (1998).
    • (1998) Science , vol.279 , pp. 1041
    • Hwang, L.H.1
  • 42
    • 0031730097 scopus 로고    scopus 로고
    • DN. Each comparison was done on four arrays composed of two pair of hybridizations done with reversal of the fluorophore labelings to eliminate biases of fluorophore incorporation. We used two different sources of mRNAs for labeled cDNA production, including a total RNA protocol (http://www.nhgri.nih.gov/DIR/LCG/15K/HTML/) and twice-purified polyadenylated mRNAs [(M. J. Marton et al. Nature Med. 4, 1293 (1998)].
    • (1998) Nature Med. , vol.4 , pp. 1293
    • Marton, M.J.1
  • 43
    • 0345227037 scopus 로고    scopus 로고
    • note
    • Supported by grants from NIH, Human Frontier Science Program, and Chiron (A.W.M.); an NIH senior fellowship (T.C.N.); an NIH grant and the Herbert W. Boyer Fund (S.M.O.); and NIH grant P41-RR11823 (B.L.D. and S.F.). S.F. is an Investigator of the Howard Hughes Medical Institute. We thank L. Hartwell, I. Herskowitz, L. Huang, and J. Rine for yeast strains and plasmids; T. Geyer, Alejandro Colman-Lerner, and R. Brent for communicating unpublished results; and D. Gottschling, I. Herskowitz, L. Pillus, J. Simon, and members of the Seattle Project and the Murray laboratory for valuable discussions.


* 이 정보는 Elsevier사의 SCOPUS DB에서 KISTI가 분석하여 추출한 것입니다.