RAC regulation of actin polymerization and proliferation by a pathway distinct from Jun kinase

Science

Volumn 274, Issue 5291, 1996, Pages 1374-1376

  Joneson, Tom ^a   McDonough, Michele ^b   Bar Sagi, Dafna ^a   Van Aelst, Linda ^b  

^a STONY BROOK UNIVERSITY   (United States)

^b Watson School of Biological Sciences   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

GUANINE NUCLEOTIDE BINDING PROTEIN; MUTANT PROTEIN; PHOSPHOTRANSFERASE; PROTEIN P21; RAC PROTEIN; UNCLASSIFIED DRUG;

ACTIN POLYMERIZATION; AMINO ACID SUBSTITUTION; ANIMAL CELL; ARTICLE; CELL PROLIFERATION; CELL STRAIN COS1; CONTROLLED STUDY; ENZYME ACTIVATION; ENZYME ACTIVITY; FIBROBLAST; NONHUMAN; PRIORITY JOURNAL;

3T3 CELLS; ACTINS; ADAPTOR PROTEINS, SIGNAL TRANSDUCING; ANIMALS; CA(2+)-CALMODULIN DEPENDENT PROTEIN KINASE; CARRIER PROTEINS; CELL DIVISION; CELL LINE; CELL LINE, TRANSFORMED; CELL MEMBRANE; COS CELLS; ENZYME ACTIVATION; GTP-BINDING PROTEINS; HUMANS; JNK MITOGEN-ACTIVATED PROTEIN KINASES; MICE; MITOGEN-ACTIVATED PROTEIN KINASES; MUTAGENESIS; PROTEIN-SERINE-THREONINE KINASES; RAC GTP-BINDING PROTEINS; RATS; TRANSFECTION;

ANIMALIA;

EID: 0029802561     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.274.5291.1374     Document Type: Article

References (39)

1. A. J. Ridley et al., Cell 70, 401 (1992).
2. O. A. Coso et al., ibid. 81, 1137 (1995); A. Minden, A. Lin, F.-X. Xavier, A. Abo, M. Karin, ibid., p. 1147; C. S. Hill, J. Wynne, R. Treisman, ibid., p. 1159.
3. O. A. Coso et al., ibid. 81, 1137 (1995); A. Minden, A. Lin, F.-X. Xavier, A. Abo, M. Karin, ibid., p. 1147; C. S. Hill, J. Wynne, R. Treisman, ibid., p. 1159.
4. O. A. Coso et al., ibid. 81, 1137 (1995); A. Minden, A. Lin, F.-X. Xavier, A. Abo, M. Karin, ibid., p. 1147; C. S. Hill, J. Wynne, R. Treisman, ibid., p. 1159.
5. R.-G. Qiu, J. Chen, D. Kim, F. McCormick, M. Symons, Nature 374, 457 (1995); R. Khosravi-Far et al., Mol. Cell. Biol. 15, 6443 (1995).
6. R.-G. Qiu, J. Chen, D. Kim, F. McCormick, M. Symons, Nature 374, 457 (1995); R. Khosravi-Far et al., Mol. Cell. Biol. 15, 6443 (1995).
7. R. Khosravi-Far et al., Mol. Cell. Biol. 16, 3923 (1996).
8. M. F. Olson, A. Ashworth, A. Hall, Science 269, 1270 (1995).
9. E. Manser et al., Nature 367, 40 (1994); U. G. Knaus S. Morris, H.-J. Dong, J. Chernoff, G. M. Bokoch, Science 269, 221 (1995); S. Bagrodia et al., J. Biol. Chem. 270, 22731 (1995); G. A. Martin et al., EMBO J. 14, 1970 (1995).
10. E. Manser et al., Nature 367, 40 (1994); U. G. Knaus S. Morris, H.-J. Dong, J. Chernoff, G. M. Bokoch, Science 269, 221 (1995); S. Bagrodia et al., J. Biol. Chem. 270, 22731 (1995); G. A. Martin et al., EMBO J. 14, 1970 (1995).
11. E. Manser et al., Nature 367, 40 (1994); U. G. Knaus S. Morris, H.-J. Dong, J. Chernoff, G. M. Bokoch, Science 269, 221 (1995); S. Bagrodia et al., J. Biol. Chem. 270, 22731 (1995); G. A. Martin et al., EMBO J. 14, 1970 (1995).
12. E. Manser et al., Nature 367, 40 (1994); U. G. Knaus S. Morris, H.-J. Dong, J. Chernoff, G. M. Bokoch, Science 269, 221 (1995); S. Bagrodia et al., J. Biol. Chem. 270, 22731 (1995); G. A. Martin et al., EMBO J. 14, 1970 (1995).
13. S. Zhang et al., J. Biol. Chem. 270, 23934 (1995); S. Bagrodia et al., ibid., p. 1; J. L Brown et al., Curr. Biol. 6, 598 (1996).
14. S. Zhang et al., J. Biol. Chem. 270, 23934 (1995); S. Bagrodia et al., ibid., p. 1; J. L Brown et al., Curr. Biol. 6, 598 (1996).
15. S. Zhang et al., J. Biol. Chem. 270, 23934 (1995); S. Bagrodia et al., ibid., p. 1; J. L Brown et al., Curr. Biol. 6, 598 (1996).
16. L. Van Aelst, T. Joneson, D. Bar-Sagi, EMBO J. 15, 3778 (1996).
17. For RAC mutant library construction and screening, full-length human RAC1 encoding a protein containing the activating G12V mutation (vaiine is substituted for glycine at position 12) was randomly mutagenized with PCR (20) and ligated in the vector pLexVJII (8) to create in-frame fusions with the LBD. Screening for mutations that affected binding of the LBD-RAC fusions to murine PAK3 or POR1 fused to the GAL4 activation domain vector pGAD1318 (8) was performed in the yeast strain L40 (8). Transformants from each screen, selected on synthetic media lacking leucine and tryptophan, were assayed for p-galactosidase activity on filters (21).
18. T. Joneson, M. McDonough, D. Bar-Sagi, L. Van Aelst, unpublished results.
19. 32P]ATP with MBP (0.2 mg/ml) as substrate. The reaction products were analyzed by SDS-polyacrylamide gel electrophoresis and visualized by autoradiography.
20. 2-terminal of c-Jun (3 μg per reaction) in kinase assay buffer.
21. 4] was microinjected into cell nuclei. For monitoring membrane ruffles and subcellular localization, cells on cover slips were fixed in 3.7% formaldehyde in phosphate-buffered saline (PBS) for 1 hour at room temperature and then permeabilized with 0.1% Triton X-100 in PBS for 3 min at room temperature. The cover slips were incubated for 1 hour at 37°C with mouse antibody to T7 epitope (Novogen) in PBS containing albumin (2 mg/mi) and then with a mixture of fluorescein-conjugated goat antibody to mouse immunoglobulin G and rhodamine-labeled phalloidin (0.01 mg/ml) (Molecular Probes). The cells were photographed with a Zeiss Axiophot fluorescence microscope.
22. T. L. Leung, E. Manser, L. Tan, L. Lim, J. Biol. Chem. 270, 29051 (1995); T. Matsui et al., EMBO J. 15, 2208 (1996).
23. T. L. Leung, E. Manser, L. Tan, L. Lim, J. Biol. Chem. 270, 29051 (1995); T. Matsui et al., EMBO J. 15, 2208 (1996).
24. K. Kimura et al., Science 273, 245 (1996).
25. L. Van Aelst, M. A. White, M. H. Wigler, Cold Spring Harbor Symp. Quant. Biol. 59, 181 (1994); M. W. White et al., Cell 80, 533 (1995).
26. L. Van Aelst, M. A. White, M. H. Wigler, Cold Spring Harbor Symp. Quant. Biol. 59, 181 (1994); M. W. White et al., Cell 80, 533 (1995).
27. T. Joneson, M. A. White, M. H. Wigler, D. Bar-Sagi, Science 271, 810 (1996).
28. M. Hibi et al., Genes Dev. 7, 2135 (1993); B. Derijard et al., Cell 76, 1025 (1994); S. Gupta, D. Campbell, B. Dérijard, R. J. Davis, Science 267, 389 (1995); H. A.-M. Abdel-Hafiz et al., Mol. Endocrinol. 6, 2079 (1992); A. J. Whitmarsh et al., Science 269, 403 (1995); H. Gille et al., Curr. Biol. 5, 1191 (1995).
29. M. Hibi et al., Genes Dev. 7, 2135 (1993); B. Derijard et al., Cell 76, 1025 (1994); S. Gupta, D. Campbell, B. Dérijard, R. J. Davis, Science 267, 389 (1995); H. A.-M. Abdel-Hafiz et al., Mol. Endocrinol. 6, 2079 (1992); A. J. Whitmarsh et al., Science 269, 403 (1995); H. Gille et al., Curr. Biol. 5, 1191 (1995).
30. M. Hibi et al., Genes Dev. 7, 2135 (1993); B. Derijard et al., Cell 76, 1025 (1994); S. Gupta, D. Campbell, B. Dérijard, R. J. Davis, Science 267, 389 (1995); H. A.-M. Abdel-Hafiz et al., Mol. Endocrinol. 6, 2079 (1992); A. J. Whitmarsh et al., Science 269, 403 (1995); H. Gille et al., Curr. Biol. 5, 1191 (1995).
31. M. Hibi et al., Genes Dev. 7, 2135 (1993); B. Derijard et al., Cell 76, 1025 (1994); S. Gupta, D. Campbell, B. Dérijard, R. J. Davis, Science 267, 389 (1995); H. A.-M. Abdel-Hafiz et al., Mol. Endocrinol. 6, 2079 (1992); A. J. Whitmarsh et al., Science 269, 403 (1995); H. Gille et al., Curr. Biol. 5, 1191 (1995).
32. M. Hibi et al., Genes Dev. 7, 2135 (1993); B. Derijard et al., Cell 76, 1025 (1994); S. Gupta, D. Campbell, B. Dérijard, R. J. Davis, Science 267, 389 (1995); H. A.-M. Abdel-Hafiz et al., Mol. Endocrinol. 6, 2079 (1992); A. J. Whitmarsh et al., Science 269, 403 (1995); H. Gille et al., Curr. Biol. 5, 1191 (1995).
33. M. Hibi et al., Genes Dev. 7, 2135 (1993); B. Derijard et al., Cell 76, 1025 (1994); S. Gupta, D. Campbell, B. Dérijard, R. J. Davis, Science 267, 389 (1995); H. A.-M. Abdel-Hafiz et al., Mol. Endocrinol. 6, 2079 (1992); A. J. Whitmarsh et al., Science 269, 403 (1995); H. Gille et al., Curr. Biol. 5, 1191 (1995).
34. M. Karin and T. Hunter, Curr. Biol. 5, 747 (1995).
35. Y. Zhou, X. Zhang, R. H. Ebright, Nucleic Acids Res. 19, 6052 (1991).
36. L. Van Aelst et al., Proc. Natl. Acad. Sci. U.S.A. 90, 6213 (1993).
37. V12L37, respectively. RAF-CAAX was cloned in the mammalian expression vector pcDNA3 (Invitrogen) and was a gift from J. Stolarov. pGAD-PAK3 was obtained by subcloning a Bam HI fragment of pcDNA3-PAK3 (gift from S. Bagrodia and R. Cerione) into pGAD1318.
38. 4 method (16) with the indicated expression plasmids (100 ng per 10-cm dish for each plasmid). Transfected cells were grown in media containing calf serum (5%) for 14 days, then fixed in 10% formaldehyde and stained with Giemsa. Foci of transformed cells appear as diffuse, darkly staining spots.
39. We thank R. Packer for technical help and J. Stolarov, S. Bagrodia, R. Cerione, and J. Chernoff for providing plasmids. Supported by Cold Spring Harbor Laboratory interim research support funds to L.V.A. and NIH grant CA55360 to D.B.-S.