TAB1 : An activator of the TAK1 MAPKKK in TGF-β signal transduction

Science

Volumn 272, Issue 5265, 1996, Pages 1179-1182

  Shibuya, Hiroshi ^a,b   Yamaguchi, Kyoko ^c   Shirakabe, Kyoko ^d   Tonegawa, Akane ^a   Gotoh, Yukiko ^d   Ueno, Naoto ^a   Irie, Kenji ^c   Nishida, Eisuke ^d   Matsumoto, Kunihiro ^c  

^a HOKKAIDO UNIVERSITY   (Japan)

^b JAPAN SCIENCE AND TECHNOLOGY AGENCY   (Japan)

^c NAGOYA UNIVERSITY   (Japan)

^d KYOTO UNIVERSITY   (Japan)

Author keywords

[No Author keywords available]

Indexed keywords

ENZYME ACTIVATOR; MITOGEN ACTIVATED PROTEIN KINASE; PHOSPHOTRANSFERASE; PLASMINOGEN ACTIVATOR INHIBITOR 1; TRANSFORMING GROWTH FACTOR BETA;

ARTICLE; CONTROLLED STUDY; ENZYME ACTIVATION; GENE ACTIVATION; HUMAN; IMMUNOPRECIPITATION; NONHUMAN; PRIORITY JOURNAL; PROMOTER REGION; PROTEIN BINDING; PROTEIN ISOLATION; PROTEIN PROTEIN INTERACTION; SIGNAL TRANSDUCTION;

ADAPTOR PROTEINS, SIGNAL TRANSDUCING; AMINO ACID SEQUENCE; ANIMALS; BASE SEQUENCE; CARRIER PROTEINS; CELL LINE; CLONING, MOLECULAR; ENZYME ACTIVATION; GENES, REPORTER; HUMANS; INTRACELLULAR SIGNALING PEPTIDES AND PROTEINS; MAP KINASE KINASE KINASES; MICE; MOLECULAR SEQUENCE DATA; PLASMINOGEN ACTIVATOR INHIBITOR 1; PROMOTER REGIONS (GENETICS); PROTEIN-SERINE-THREONINE KINASES; RECOMBINANT FUSION PROTEINS; SACCHAROMYCES CEREVISIAE; SIGNAL TRANSDUCTION; TRANSFECTION; TRANSFORMATION, GENETIC; TRANSFORMING GROWTH FACTOR BETA;

EID: 0029940355     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.272.5265.1179     Document Type: Article

References (31)

1. E. Nishida and Y. Gotoh, Trends Biochem. Sci. 18, 128 (1993); K. J. Blumer and G. L. Johnson, ibid. 19, 236 (1994); R. J. Davis, ibid., p. 470; C. J. Marshall, Cell 80, 179 (1995).
2. E. Nishida and Y. Gotoh, Trends Biochem. Sci. 18, 128 (1993); K. J. Blumer and G. L. Johnson, ibid. 19, 236 (1994); R. J. Davis, ibid., p. 470; C. J. Marshall, Cell 80, 179 (1995).
3. E. Nishida and Y. Gotoh, Trends Biochem. Sci. 18, 128 (1993); K. J. Blumer and G. L. Johnson, ibid. 19, 236 (1994); R. J. Davis, ibid., p. 470; C. J. Marshall, Cell 80, 179 (1995).
4. E. Nishida and Y. Gotoh, Trends Biochem. Sci. 18, 128 (1993); K. J. Blumer and G. L. Johnson, ibid. 19, 236 (1994); R. J. Davis, ibid., p. 470; C. J. Marshall, Cell 80, 179 (1995).
5. K. Yamaguchi et al., Science 270, 2008 (1995).
6. J. L. Wrana, L. Attisano, R. Wieser, F. Ventura, J. Massague, Nature 370, 341 (1994); D. M. Kingsley, Genes Dev. 8, 133 (1994).
7. J. L. Wrana, L. Attisano, R. Wieser, F. Ventura, J. Massague, Nature 370, 341 (1994); D. M. Kingsley, Genes Dev. 8, 133 (1994).
8. S. Felds and R. Sternglanz, Trends Genet. 10, 285 (1994).
9. The pLexA-TAK1ΔN plasmid comprises the TAK1ΔN coding sequence (TAK1 residues 21 to 579) (2) inserted in frame into pBTM116 [A. B. Vojtek, S. M. Hollenberg, J. A. Cooper, Cell 74, 205 (1993)]. The yeast two-hybrid system was used to identify proteins encoded by a human brain cDNA library that interact with TAK1ΔN. The two hybrids were expressed in a Saccharamyces cerevisiae strain L40 (LYS2::lexA-HIS3) that contains an integrated reporter construct in which a binding site for the LexA protein was placed upstream of the yeast HIS3 coding region. If the two hybrid proteins interact, then transactivation of the reporter construct occurs and the yeast cells can grow in the absence of histidine (SC-His). The LexA-TAK1ΔN fusion protein alone induced sufficient HIS3 expression to allow growth in the absence of exogenous histidine. However, histidine auxotrophy could be reestablished by growing cells in the presence of 40 mM 3-aminotriozole, a chemical inhibitor of the HIS3 product, imidazole glycerol dehydrogenase [G. M. Kishore and D. M. Shah, Annu Rev. Biochem. 57, 627 (1988)].
10. The pLexA-TAK1ΔN plasmid comprises the TAK1ΔN coding sequence (TAK1 residues 21 to 579) (2) inserted in frame into pBTM116 [A. B. Vojtek, S. M. Hollenberg, J. A. Cooper, Cell 74, 205 (1993)]. The yeast two-hybrid system was used to identify proteins encoded by a human brain cDNA library that interact with TAK1ΔN. The two hybrids were expressed in a Saccharamyces cerevisiae strain L40 (LYS2::lexA-HIS3) that contains an integrated reporter construct in which a binding site for the LexA protein was placed upstream of the yeast HIS3 coding region. If the two hybrid proteins interact, then transactivation of the reporter construct occurs and the yeast cells can grow in the absence of histidine (SC-His). The LexA-TAK1ΔN fusion protein alone induced sufficient HIS3 expression to allow growth in the absence of exogenous histidine. However, histidine auxotrophy could be reestablished by growing cells in the presence of 40 mM 3-aminotriozole, a chemical inhibitor of the HIS3 product, imidazole glycerol dehydrogenase [G. M. Kishore and D. M. Shah, Annu Rev. Biochem. 57, 627 (1988)].
11. K. Irie et al., Science 265, 1716 (1994).
12. p368-dependent manner.
13. p368-dependent manner.
14. p368-dependent manner.
15. The DNA sequence that encodes the HA epitope recognized by the monoclonal antibody 12CA5 was attached in frame to the DNA encoding the COOH termini of TAK1 and TAK1 (K63W) by the polymerase chain reaction (PCR). All constructs were expressed from the TDH3 promoter. The TAB1 expression plasmid, pGAP-HTH9M, encodes the COOH-terminal 68 amino acids of TAB1. The coding sequence for the COOH-terminal 68 amino acids of TAB1 was amplified by PCR with a 5′ primer (5′-GAGAATTCATGCGGCAAAGC-3′) incorporating an Eco RI site and an ATG codon, and a 3′ primer (5′-GGGTCGACTACGGTGC-3′) incorporating a Sal I site. A 240-base pair Eco RI-Sal I fragment generated by PCR was inserted into the Eco RI-Sal I gap of YEp-GAP112, a multicopy TRP1 plasmid that contains the TDH3 promoter [H. Banno et al., Mol. Cell. Biol. 13, 4745 (1993)], to generate pGAP-HTH9M.
16. 2,-terminal end of the coding sequence corresponds to the Kozak consensus [M. Kozak, J. Cell Biol. 108, 229 (1989)], and there are no upstream ATG codons.
17. S. Ohta et al., FEBS Lett. 314, 356 (1992).
18. 2-terminus of TAB1. The pCS2MT-TAB1 plasmid was digested with Bam HI and Xba I, and the resulting fragment was isolated and inserted into the Eco Rl-Xba I sites of the mammalian expression vector pEF, in which expression is controlled by the human elongation factor 1α (EF1α) gene promoter.
19. M. R. Keeton, S. A. Curriden, A.-J. Zonneveld, D. J. Loskutoff, J. Biol. Chem. 266, 23048 (1991).
20. M. Abe et al. Anal. Biochem. 216, 276 (1994).
21. The TAB1 expression plasmid, pEF-TAB1, contains the full-length TAB1 coding sequence under the control of the EF1α gene promoter. This plasrnid was generated by cleaving the pEF vector with Eco RI and inserting the Eco RI fragment from pBS-TAB1, which itself was generated by subclonnig of the TAB1 cDNA into the Eco RI site of pBS.
22. K. Yamaguchi, K. Irie, K. Matsumoto, unpublished data.
23. Abbreviations for the amino acid residues: 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.
24. Yeast strain L40 was cotransformed with (i) an expression vector encoding full-length TAK1 or a deletion construct fused to the LexA DNA binding domain and (ii) pGAD-TAB1 or pGAD-TAB2. Interaction between fusion proteins expressed by the specified plasmids was demonstrated by the ability of the yeast cells to grow on plates of SC-Hs containing 40 mM 3-aminotriozole. Each deletion construct of TAK1 was generated from the full-length TAK1.
25. P368 allele under the control of the CVC1 promoter (6). Plasmids pNV11-HU11 and pNV11-HU11F express, under the control of the TDH3 promoter, TAK1ΔN and full-length TAK1, respectively (2).
26. Yeast cells (60-ml culture) were grown to an optical density at 600 nm of 0.8 Cell extracts were prepared with a lysis buffer as described (6) and centrifuged for 30 min at 100,000g. The supernatant was subjected to immunoprecipitation with antibodies to HA. Briefly, a portion (300 μl) of the supernatant was mixed with 2 μl of antibodies and 90 μl of protein A-Sepharose (Pharmacia), and the immune complex was washed three times with lysis buffer and assayed for kinase activity (2). Immunoblot analysis of each immunoprecipitate with monoclonal antibody 12CA5 to HA demonstrated that approximately the same amount of TAK1-HA or TAK1(K63W)-HA was recovered in each sample, indicating that TAB1 expression did not affect the amount of TAK1 expressed.
27. H. Shibuya et al., Nature 357, 700 (1992).
28. H. Shibuya et al., Mol. Cell. Biol. 14, 5812 (1994).
29. Transfection efficiency was normalized by cotransfection with the pXeX-β-Gal vector [A. D. Johnson and P. A. Krieg, Gene 147, 223 (1994)] in all luciferase reporter experiments. The β-galatosidase assay was performed as described by the manufacturer (Clontech) with the cell lysates prepared for luciferase assay.
30. A 1.3-kb Eco RI-Hinc II fragment of pBS-TAB1 (14) that encodes amino acids 1 to 418 of TAB1 was subcloned into the pKT10 vector to generate pKT10-TAB1(1-418). The pEF-TAB1(1-418) plasmid was generated by cleaving the pEF vector with Eco RI and Sal I and inserting the Eco RI-Sal I fragment from pKT10-TAB1(1-418),
31. We thank M. Abe, B. Errede, A. Johnson, and D. Turner for materials; R. Ruggieri and R. Yu for critical reading of the manuscript; and H. Wang and J. Reed for helpful discussions and sharing data before publication. Supported by a special grant for Advanced Research on Cancer from the Ministry of Education, Culture, and Science of Japan (Io K.M.).