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Structural Requirements for PAK Activation by Rac GTPases

Knaus, Ulla G., Wang, Yan, Reilly, Abina M., Warnock, Dawn, Jackson, Janis H.·1998·Journal of Biological Chemistry
DOI 10.1074/jbc.273.34.21512
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FIG. 1 The polybasic domain of Rac1 conveys high binding affinity to PAK1. Decreasing amounts of lysate (15, 10, 5, 2.5, and 1.25 m l) from PAK1(wt)-overexpressing COS-7 cells were electrophoresed and blotted onto nitrocellulose. After renaturation, the blots were probed with 1 m g/ml [ 35 S]GTP g S-labeled wild type and mutant Rac proteins as indicated. The autoradiograms of the blots were quantitated by a PhosphorImager. The depicted values are representative of three separate experiments. (Rac mutants are listed in Table I).
FIG. 1 The polybasic domain of Rac1 conveys high binding affinity to PAK1. Decreasing amounts of lysate (15, 10, 5, 2.5, and 1.25 m l) from PAK1(wt)-overexpressing COS-7 cells were electrophoresed and blotted onto nitrocellulose. After renaturation, the blots were probed with 1 m g/ml [ 35 S]GTP g S-labeled wild type and mutant Rac proteins as indicated. The autoradiograms of the blots were quantitated by a PhosphorImager. The depicted values are representative of three separate experiments. (Rac mutants are listed in Table I).CC-BY · From the PDF
PAK 1 (wt)
PAK 1 (wt)CC-BY · From the PDF
FIG. 3 The polybasic domain of Rac1 is required for PAK1induced phosphorylation of exogenous substrates. Immunoprecipitated wild type PAK1 from COS-7 cell lysates was incubated with 1 m g of GTP g S-labeled wild type and mutant Rac proteins ( E. coli -derived GTPases) as indicated, along with the addition of a peptide derived from the NADPH oxidase component p47 phox . Peptide phosphorylation was assessed on phosphocellulose filters, which were subjected to scintillation counting. Control values representing PAK autophosphorylation by itself were subtracted. Final results are expressed as the mean percentage 6 S.D. of four independent experiments, with PAK1 activity induced by Rac1(wt) set at 100%.
FIG. 3 The polybasic domain of Rac1 is required for PAK1induced phosphorylation of exogenous substrates. Immunoprecipitated wild type PAK1 from COS-7 cell lysates was incubated with 1 m g of GTP g S-labeled wild type and mutant Rac proteins ( E. coli -derived GTPases) as indicated, along with the addition of a peptide derived from the NADPH oxidase component p47 phox . Peptide phosphorylation was assessed on phosphocellulose filters, which were subjected to scintillation counting. Control values representing PAK autophosphorylation by itself were subtracted. Final results are expressed as the mean percentage 6 S.D. of four independent experiments, with PAK1 activity induced by Rac1(wt) set at 100%.CC-BY · From the PDF
FIG. 4 The polybasic domain of Rac1 is required for efficient PAK1 activation in vivo . Wild type PAK1 was co-expressed for 24 h with activated forms of Rac1, Rac2, and the polybasic domain mutant Rac1(6Q) (see Table I) in HeLa cells (DNA ratio 9:1). Stimulation of PAK activity was visualized by anti-Myc immunoblots of cell lysates and quantitated by a PhosphorImager. Activation of PAK1 (68 kDa) results in autophosphorylated, slower migrating bands on 6.5% gels ( panel 1 ). The expression levels of the activated Rac proteins in the same lysates were confirmed with anti-Myc immunoblots ( panel 2 ) and subsequently quantitated. Similar results were obtained in two independent experiments.
FIG. 4 The polybasic domain of Rac1 is required for efficient PAK1 activation in vivo . Wild type PAK1 was co-expressed for 24 h with activated forms of Rac1, Rac2, and the polybasic domain mutant Rac1(6Q) (see Table I) in HeLa cells (DNA ratio 9:1). Stimulation of PAK activity was visualized by anti-Myc immunoblots of cell lysates and quantitated by a PhosphorImager. Activation of PAK1 (68 kDa) results in autophosphorylated, slower migrating bands on 6.5% gels ( panel 1 ). The expression levels of the activated Rac proteins in the same lysates were confirmed with anti-Myc immunoblots ( panel 2 ) and subsequently quantitated. Similar results were obtained in two independent experiments.CC-BY · From the PDF
FIG. 5 The polybasic domain of PAK1 is required for efficient activation by GTPases. A , PAK1(wt) and mutant PAK1(K66-68L), PAK1(K66-68R), and PAK1(K66-68Q) were expressed in COS-7 cells, blotted in expression level-adjusted amounts, and compared for GTPRac1, -Rac2, and -Cdc42 binding with overlay assays. The mean values 6 S.D. of three independent experiments are shown. The binding of each GTPase to PAK1(wt) is normalized to 100% for easier comparison within one group. B , wild type PAK1 or the PAK1 mutants PAK1(K6668L), PAK1(K66-68R), or PAK1(K66-68Q) were co-transfected with activated forms of Rac1, Rac2, and Cdc42 in HeLa cells, as described under 'Experimental Procedures' (DNA ratio 4:1), and cell lysates were prepared after 24 h. PAK activity was assessed by Western blotting with anti-Myc antibody, detecting the faster migrating, nonactivated and the slower migrating, hyperphosphorylated active forms of PAK1. Rac1, Rac2, and Cdc42 expression levels were monitored in the same cell lysates with anti-Myc antibody. The PAK1(wt) activation by each GTPase was normalized to 100% as described for A . The bar graph shows one of two similar experiments.
FIG. 5 The polybasic domain of PAK1 is required for efficient activation by GTPases. A , PAK1(wt) and mutant PAK1(K66-68L), PAK1(K66-68R), and PAK1(K66-68Q) were expressed in COS-7 cells, blotted in expression level-adjusted amounts, and compared for GTPRac1, -Rac2, and -Cdc42 binding with overlay assays. The mean values 6 S.D. of three independent experiments are shown. The binding of each GTPase to PAK1(wt) is normalized to 100% for easier comparison within one group. B , wild type PAK1 or the PAK1 mutants PAK1(K6668L), PAK1(K66-68R), or PAK1(K66-68Q) were co-transfected with activated forms of Rac1, Rac2, and Cdc42 in HeLa cells, as described under 'Experimental Procedures' (DNA ratio 4:1), and cell lysates were prepared after 24 h. PAK activity was assessed by Western blotting with anti-Myc antibody, detecting the faster migrating, nonactivated and the slower migrating, hyperphosphorylated active forms of PAK1. Rac1, Rac2, and Cdc42 expression levels were monitored in the same cell lysates with anti-Myc antibody. The PAK1(wt) activation by each GTPase was normalized to 100% as described for A . The bar graph shows one of two similar experiments.CC-BY · From the PDF

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Research works printed but not linked: 44

  1. Hall, 1994, Annu. Rev. Cell Biol., 10, 31, 10.1146/annurev.cb.10.110194.000335 · DOI
  2. Ridley, 1992, Cell, 70, 389, 10.1016/0092-8674(92)90163-7 · DOI
  3. Ridley, 1992, Cell, 70, 401, 10.1016/0092-8674(92)90164-8 · DOI
  4. Kozma, 1995, Mol. Cell. Biol., 16, 5069, 10.1128/MCB.16.9.5069 · DOI
  5. Nobes, 1995, Cell, 81, 53, 10.1016/0092-8674(95)90370-4 · DOI
  6. Coso, 1995, Cell, 81, 1137, 10.1016/S0092-8674(05)80018-2 · DOI · Paper page
  7. Minden, 1995, Cell, 81, 1147, 10.1016/S0092-8674(05)80019-4 · DOI
  8. Olson, 1995, Science, 269, 1270, 10.1126/science.7652575 · DOI
  9. Zhang, 1995, J. Biol. Chem., 270, 23934, 10.1074/jbc.270.41.23934 · DOI · Paper page
  10. Bagrodia, 1995, J. Biol. Chem., 270, 27995, 10.1074/jbc.270.47.27995 · DOI
  11. Hill, 1995, Cell, 81, 1159, 10.1016/S0092-8674(05)80020-0 · DOI
  12. Qiu, 1995, Nature, 374, 457, 10.1038/374457a0 · DOI
  13. Khosravi-Far, 1995, Mol. Cell. Biol., 15, 6443, 10.1128/MCB.15.11.6443 · DOI · Paper page
  14. Abo, 1991, Nature, 353, 668, 10.1038/353668a0 · DOI
  15. Knaus, 1991, Science, 254, 1512, 10.1126/science.1660188 · DOI · Paper page
  16. Heyworth, 1993, Mol. Biol. Cell, 4, 261, 10.1091/mbc.4.3.261 · DOI
  17. Dorseuil, 1996, J. Biol. Chem., 271, 83, 10.1074/jbc.271.1.83 · DOI
  18. Manser, 1994, Nature, 367, 40, 10.1038/367040a0 · DOI
  19. Polverino, 1995, J. Biol. Chem., 270, 26067, 10.1074/jbc.270.44.26067 · DOI
  20. Dharmawardhane, 1997, J. Cell Biol., 138, 1265, 10.1083/jcb.138.6.1265 · DOI
  21. Sells, 1997, Curr. Biol., 7, 202, 10.1016/S0960-9822(97)70091-5 · DOI · Paper page
  22. Brzeska, 1997, Proc. Natl. Acad. Sci. U. S. A., 94, 1092, 10.1073/pnas.94.4.1092 · DOI
  23. Wu, 1996, J. Biol. Chem., 271, 31787, 10.1074/jbc.271.50.31787 · DOI
  24. Tauzon, 1984, J. Biol. Chem., 259, 541, 10.1016/S0021-9258(17)43695-7 · DOI
  25. Knaus, 1995, Science, 269, 221, 10.1126/science.7618083 · DOI
  26. Jackson, 1990, Proc. Natl. Acad. Sci. U. S. A., 87, 3042, 10.1073/pnas.87.8.3042 · DOI
  27. Knaus, 1992, J. Biol. Chem., 267, 23575, 10.1016/S0021-9258(18)35877-0 · DOI
  28. Burbelo, 1995, J. Biol. Chem., 270, 29071, 10.1074/jbc.270.49.29071 · DOI
  29. Symons, 1996, Cell, 84, 723, 10.1016/S0092-8674(00)81050-8 · DOI
  30. Sells, 1997, Trends Cell Biol., 7, 162, 10.1016/S0962-8924(97)01003-9 · DOI
  31. Bagrodia, 1995, J. Biol. Chem., 270, 22731, 10.1074/jbc.270.39.22731 · DOI
  32. Bokoch, 1996, J. Biol. Chem., 271, 25746, 10.1074/jbc.271.42.25746 · DOI
  33. Lamarche, 1996, Cell, 87, 519, 10.1016/S0092-8674(00)81371-9 · DOI
  34. Westwick, 1997, Mol. Cell. Biol., 17, 1324, 10.1128/MCB.17.3.1324 · DOI
  35. Diekmann, 1995, EMBO J., 14, 5297, 10.1002/j.1460-2075.1995.tb00214.x · DOI · Paper page
  36. Hancock, 1990, Cell, 63, 133, 10.1016/0092-8674(90)90294-O · DOI
  37. Kreck, 1994, J. Biol. Chem., 269, 4161, 10.1016/S0021-9258(17)41757-1 · DOI · Paper page
  38. Kreck, 1996, Biochemistry, 35, 15683, 10.1021/bi962064l · DOI
  39. Joseph, 1995, J. Biol. Chem., 270, 29079, 10.1074/jbc.270.49.29079 · DOI
  40. Jackson, 1994, Proc. Natl. Acad. Sci. U. S. A., 91, 12730, 10.1073/pnas.91.26.12730 · DOI
  41. Shirataki, 1991, J. Biol. Chem., 266, 20672, 10.1016/S0021-9258(18)54761-X · DOI
  42. Manser, 1994, Nature, 363, 364, 10.1038/363364a0 · DOI
  43. Manser, 1995, J. Biol. Chem., 270, 25070, 10.1074/jbc.270.42.25070 · DOI
  44. Manser, 1998, Mol. Cell, 1, 183, 10.1016/S1097-2765(00)80019-2 · DOI

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Metadata

Authors
Knaus, U. G., Wang, Y., Reilly, A. M., Warnock, D., & Jackson, J. H.
Venue
Journal of Biological Chemistry
Publisher
Elsevier BV
Published
1998-08
Volume
273
Issue
34
Pages
21512-21518
ISSN
0021-9258
DOI
10.1074/jbc.273.34.21512
Cited by
111
References
0
Language
en
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