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. 1995 Feb;15(2):1110–1122. doi: 10.1128/mcb.15.2.1110

Regulation of scatter factor/hepatocyte growth factor responses by Ras, Rac, and Rho in MDCK cells.

A J Ridley 1, P M Comoglio 1, A Hall 1
PMCID: PMC232019  PMID: 7823927

Abstract

Scatter factor/hepatocyte growth factor (SF/HGF) stimulates the motility of epithelial cells, initially inducing centrifugal spreading of cell colonies followed by disruption of cell-cell junctions and subsequent cell scattering. These responses are accompanied by changes in the actin cytoskeleton, including increased membrane ruffling and lamellipodium extension, disappearance of peripheral actin bundles at the edges of colonies, and an overall decrease in stress fibers. The roles of the small GTP-binding proteins Ras, Rac, and Rho in regulating responses to SF/HGF were investigated by microinjection. Inhibition of endogenous Ras proteins prevented SF/HGF-induced actin reorganization, spreading, and scattering, whereas microinjection of activated H-Ras protein stimulated spreading and actin reorganization but not scattering. When a dominant inhibitor of Rac was injected, SF/HGF- and Ras-induced spreading and actin reorganization were prevented, although activated Rac alone did not stimulate either response. Microinjection of activated Rho inhibited spreading and scattering, while inhibition of Rho function led to the disappearance of stress fibers and peripheral bundles but did not prevent SF/HGF-induced motility. We conclude that Ras and Rac act downstream of the SF/HGF receptor p190Met to mediate cell spreading but that an additional signal is required to induce scattering.

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Selected References

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  1. Aktories K., Braun U., Rösener S., Just I., Hall A. The rho gene product expressed in E. coli is a substrate of botulinum ADP-ribosyltransferase C3. Biochem Biophys Res Commun. 1989 Jan 16;158(1):209–213. doi: 10.1016/s0006-291x(89)80199-8. [DOI] [PubMed] [Google Scholar]
  2. Bar-Sagi D., Feramisco J. R. Induction of membrane ruffling and fluid-phase pinocytosis in quiescent fibroblasts by ras proteins. Science. 1986 Sep 5;233(4768):1061–1068. doi: 10.1126/science.3090687. [DOI] [PubMed] [Google Scholar]
  3. Bardelli A., Maina F., Gout I., Fry M. J., Waterfield M. D., Comoglio P. M., Ponzetto C. Autophosphorylation promotes complex formation of recombinant hepatocyte growth factor receptor with cytoplasmic effectors containing SH2 domains. Oncogene. 1992 Oct;7(10):1973–1978. [PubMed] [Google Scholar]
  4. Behrens J., Mareel M. M., Van Roy F. M., Birchmeier W. Dissecting tumor cell invasion: epithelial cells acquire invasive properties after the loss of uvomorulin-mediated cell-cell adhesion. J Cell Biol. 1989 Jun;108(6):2435–2447. doi: 10.1083/jcb.108.6.2435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bement W. M., Forscher P., Mooseker M. S. A novel cytoskeletal structure involved in purse string wound closure and cell polarity maintenance. J Cell Biol. 1993 May;121(3):565–578. doi: 10.1083/jcb.121.3.565. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bottaro D. P., Rubin J. S., Faletto D. L., Chan A. M., Kmiecik T. E., Vande Woude G. F., Aaronson S. A. Identification of the hepatocyte growth factor receptor as the c-met proto-oncogene product. Science. 1991 Feb 15;251(4995):802–804. doi: 10.1126/science.1846706. [DOI] [PubMed] [Google Scholar]
  7. Bretscher A. Microfilament structure and function in the cortical cytoskeleton. Annu Rev Cell Biol. 1991;7:337–374. doi: 10.1146/annurev.cb.07.110191.002005. [DOI] [PubMed] [Google Scholar]
  8. Burridge K., Fath K., Kelly T., Nuckolls G., Turner C. Focal adhesions: transmembrane junctions between the extracellular matrix and the cytoskeleton. Annu Rev Cell Biol. 1988;4:487–525. doi: 10.1146/annurev.cb.04.110188.002415. [DOI] [PubMed] [Google Scholar]
  9. Bussolino F., Di Renzo M. F., Ziche M., Bocchietto E., Olivero M., Naldini L., Gaudino G., Tamagnone L., Coffer A., Comoglio P. M. Hepatocyte growth factor is a potent angiogenic factor which stimulates endothelial cell motility and growth. J Cell Biol. 1992 Nov;119(3):629–641. doi: 10.1083/jcb.119.3.629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Bögler O., Entwistle A., Kuhn R., Monuki E., Lemke G., Noble M. Single cell analysis of the expression of a nuclear protein, SCIP, by fluorescent immunohistochemistry visualized with confocal microscopy. Histochem J. 1993 Oct;25(10):746–761. doi: 10.1007/BF00211770. [DOI] [PubMed] [Google Scholar]
  11. Cooper J. A. The role of actin polymerization in cell motility. Annu Rev Physiol. 1991;53:585–605. doi: 10.1146/annurev.ph.53.030191.003101. [DOI] [PubMed] [Google Scholar]
  12. Dowrick P. G., Prescott A. R., Warn R. M. Scatter factor affects major changes in the cytoskeletal organization of epithelial cells. Cytokine. 1991 Jul;3(4):299–310. doi: 10.1016/1043-4666(91)90498-3. [DOI] [PubMed] [Google Scholar]
  13. Furth M. E., Davis L. J., Fleurdelys B., Scolnick E. M. Monoclonal antibodies to the p21 products of the transforming gene of Harvey murine sarcoma virus and of the cellular ras gene family. J Virol. 1982 Jul;43(1):294–304. doi: 10.1128/jvi.43.1.294-304.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gherardi E., Gray J., Stoker M., Perryman M., Furlong R. Purification of scatter factor, a fibroblast-derived basic protein that modulates epithelial interactions and movement. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5844–5848. doi: 10.1073/pnas.86.15.5844. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gherardi E., Stoker M. Hepatocyte growth factor--scatter factor: mitogen, motogen, and met. Cancer Cells. 1991 Jun;3(6):227–232. [PubMed] [Google Scholar]
  16. Giordano S., Zhen Z., Medico E., Gaudino G., Galimi F., Comoglio P. M. Transfer of motogenic and invasive response to scatter factor/hepatocyte growth factor by transfection of human MET protooncogene. Proc Natl Acad Sci U S A. 1993 Jan 15;90(2):649–653. doi: 10.1073/pnas.90.2.649. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Grant D. S., Kleinman H. K., Goldberg I. D., Bhargava M. M., Nickoloff B. J., Kinsella J. L., Polverini P., Rosen E. M. Scatter factor induces blood vessel formation in vivo. Proc Natl Acad Sci U S A. 1993 Mar 1;90(5):1937–1941. doi: 10.1073/pnas.90.5.1937. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Graziani A., Gramaglia D., Cantley L. C., Comoglio P. M. The tyrosine-phosphorylated hepatocyte growth factor/scatter factor receptor associates with phosphatidylinositol 3-kinase. J Biol Chem. 1991 Nov 25;266(33):22087–22090. [PubMed] [Google Scholar]
  19. Graziani A., Gramaglia D., dalla Zonca P., Comoglio P. M. Hepatocyte growth factor/scatter factor stimulates the Ras-guanine nucleotide exchanger. J Biol Chem. 1993 May 5;268(13):9165–9168. [PubMed] [Google Scholar]
  20. Hall A. The cellular functions of small GTP-binding proteins. Science. 1990 Aug 10;249(4969):635–640. doi: 10.1126/science.2116664. [DOI] [PubMed] [Google Scholar]
  21. Hartmann G., Naldini L., Weidner K. M., Sachs M., Vigna E., Comoglio P. M., Birchmeier W. A functional domain in the heavy chain of scatter factor/hepatocyte growth factor binds the c-Met receptor and induces cell dissociation but not mitogenesis. Proc Natl Acad Sci U S A. 1992 Dec 1;89(23):11574–11578. doi: 10.1073/pnas.89.23.11574. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Herman I. M., Crisona N. J., Pollard T. D. Relation between cell activity and the distribution of cytoplasmic actin and myosin. J Cell Biol. 1981 Jul;90(1):84–91. doi: 10.1083/jcb.90.1.84. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hinsch K. D., Habermann B., Just I., Hinsch E., Pfisterer S., Schill W. B., Aktories K. ADP-ribosylation of Rho proteins inhibits sperm motility. FEBS Lett. 1993 Nov 8;334(1):32–36. doi: 10.1016/0014-5793(93)81674-o. [DOI] [PubMed] [Google Scholar]
  24. Kemler R. From cadherins to catenins: cytoplasmic protein interactions and regulation of cell adhesion. Trends Genet. 1993 Sep;9(9):317–321. doi: 10.1016/0168-9525(93)90250-l. [DOI] [PubMed] [Google Scholar]
  25. Leevers S. J., Marshall C. J. MAP kinase regulation--the oncogene connection. Trends Cell Biol. 1992 Oct;2(10):283–286. doi: 10.1016/0962-8924(92)90105-v. [DOI] [PubMed] [Google Scholar]
  26. Lloyd A. C., Paterson H. F., Morris J. D., Hall A., Marshall C. J. p21H-ras-induced morphological transformation and increases in c-myc expression are independent of functional protein kinase C. EMBO J. 1989 Apr;8(4):1099–1104. doi: 10.1002/j.1460-2075.1989.tb03479.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Luna E. J., Hitt A. L. Cytoskeleton--plasma membrane interactions. Science. 1992 Nov 6;258(5084):955–964. doi: 10.1126/science.1439807. [DOI] [PubMed] [Google Scholar]
  28. Martin P., Lewis J. Actin cables and epidermal movement in embryonic wound healing. Nature. 1992 Nov 12;360(6400):179–183. doi: 10.1038/360179a0. [DOI] [PubMed] [Google Scholar]
  29. Michalopoulos G., Houck K. A., Dolan M. L., Leutteke N. C. Control of hepatocyte replication by two serum factors. Cancer Res. 1984 Oct;44(10):4414–4419. [PubMed] [Google Scholar]
  30. Miyazawa K., Shimomura T., Kitamura A., Kondo J., Morimoto Y., Kitamura N. Molecular cloning and sequence analysis of the cDNA for a human serine protease reponsible for activation of hepatocyte growth factor. Structural similarity of the protease precursor to blood coagulation factor XII. J Biol Chem. 1993 May 15;268(14):10024–10028. [PubMed] [Google Scholar]
  31. Miyazawa K., Tsubouchi H., Naka D., Takahashi K., Okigaki M., Arakaki N., Nakayama H., Hirono S., Sakiyama O., Takahashi K. Molecular cloning and sequence analysis of cDNA for human hepatocyte growth factor. Biochem Biophys Res Commun. 1989 Sep 15;163(2):967–973. doi: 10.1016/0006-291x(89)92316-4. [DOI] [PubMed] [Google Scholar]
  32. Morii N., Teru-uchi T., Tominaga T., Kumagai N., Kozaki S., Ushikubi F., Narumiya S. A rho gene product in human blood platelets. II. Effects of the ADP-ribosylation by botulinum C3 ADP-ribosyltransferase on platelet aggregation. J Biol Chem. 1992 Oct 15;267(29):20921–20926. [PubMed] [Google Scholar]
  33. Nakamura T., Nawa K., Ichihara A. Partial purification and characterization of hepatocyte growth factor from serum of hepatectomized rats. Biochem Biophys Res Commun. 1984 Aug 16;122(3):1450–1459. doi: 10.1016/0006-291x(84)91253-1. [DOI] [PubMed] [Google Scholar]
  34. Nakamura T., Nishizawa T., Hagiya M., Seki T., Shimonishi M., Sugimura A., Tashiro K., Shimizu S. Molecular cloning and expression of human hepatocyte growth factor. Nature. 1989 Nov 23;342(6248):440–443. doi: 10.1038/342440a0. [DOI] [PubMed] [Google Scholar]
  35. Naldini L., Tamagnone L., Vigna E., Sachs M., Hartmann G., Birchmeier W., Daikuhara Y., Tsubouchi H., Blasi F., Comoglio P. M. Extracellular proteolytic cleavage by urokinase is required for activation of hepatocyte growth factor/scatter factor. EMBO J. 1992 Dec;11(13):4825–4833. doi: 10.1002/j.1460-2075.1992.tb05588.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Naldini L., Vigna E., Narsimhan R. P., Gaudino G., Zarnegar R., Michalopoulos G. K., Comoglio P. M. Hepatocyte growth factor (HGF) stimulates the tyrosine kinase activity of the receptor encoded by the proto-oncogene c-MET. Oncogene. 1991 Apr;6(4):501–504. [PubMed] [Google Scholar]
  37. Naldini L., Weidner K. M., Vigna E., Gaudino G., Bardelli A., Ponzetto C., Narsimhan R. P., Hartmann G., Zarnegar R., Michalopoulos G. K. Scatter factor and hepatocyte growth factor are indistinguishable ligands for the MET receptor. EMBO J. 1991 Oct;10(10):2867–2878. doi: 10.1002/j.1460-2075.1991.tb07836.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Nishiyama T., Sasaki T., Takaishi K., Kato M., Yaku H., Araki K., Matsuura Y., Takai Y. rac p21 is involved in insulin-induced membrane ruffling and rho p21 is involved in hepatocyte growth factor- and 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced membrane ruffling in KB cells. Mol Cell Biol. 1994 Apr;14(4):2447–2456. doi: 10.1128/mcb.14.4.2447. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Paterson H. F., Self A. J., Garrett M. D., Just I., Aktories K., Hall A. Microinjection of recombinant p21rho induces rapid changes in cell morphology. J Cell Biol. 1990 Sep;111(3):1001–1007. doi: 10.1083/jcb.111.3.1001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Pepper M. S., Matsumoto K., Nakamura T., Orci L., Montesano R. Hepatocyte growth factor increases urokinase-type plasminogen activator (u-PA) and u-PA receptor expression in Madin-Darby canine kidney epithelial cells. J Biol Chem. 1992 Oct 5;267(28):20493–20496. [PubMed] [Google Scholar]
  41. Ponzetto C., Bardelli A., Zhen Z., Maina F., dalla Zonca P., Giordano S., Graziani A., Panayotou G., Comoglio P. M. A multifunctional docking site mediates signaling and transformation by the hepatocyte growth factor/scatter factor receptor family. Cell. 1994 Apr 22;77(2):261–271. doi: 10.1016/0092-8674(94)90318-2. [DOI] [PubMed] [Google Scholar]
  42. Ridley A. J., Hall A. The small GTP-binding protein rho regulates the assembly of focal adhesions and actin stress fibers in response to growth factors. Cell. 1992 Aug 7;70(3):389–399. doi: 10.1016/0092-8674(92)90163-7. [DOI] [PubMed] [Google Scholar]
  43. Ridley A. J. Membrane ruffling and signal transduction. Bioessays. 1994 May;16(5):321–327. doi: 10.1002/bies.950160506. [DOI] [PubMed] [Google Scholar]
  44. Ridley A. J., Paterson H. F., Johnston C. L., Diekmann D., Hall A. The small GTP-binding protein rac regulates growth factor-induced membrane ruffling. Cell. 1992 Aug 7;70(3):401–410. doi: 10.1016/0092-8674(92)90164-8. [DOI] [PubMed] [Google Scholar]
  45. Rong S., Bodescot M., Blair D., Dunn J., Nakamura T., Mizuno K., Park M., Chan A., Aaronson S., Vande Woude G. F. Tumorigenicity of the met proto-oncogene and the gene for hepatocyte growth factor. Mol Cell Biol. 1992 Nov;12(11):5152–5158. doi: 10.1128/mcb.12.11.5152. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Rosen E. M., Meromsky L., Goldberg I., Bhargava M., Setter E. Studies on the mechanism of scatter factor. Effects of agents that modulate intracellular signal transduction, macromolecule synthesis and cytoskeleton assembly. J Cell Sci. 1990 Aug;96(Pt 4):639–649. doi: 10.1242/jcs.96.4.639. [DOI] [PubMed] [Google Scholar]
  47. Rosen E. M., Meromsky L., Setter E., Vinter D. W., Goldberg I. D. Smooth muscle-derived factor stimulates mobility of human tumor cells. Invasion Metastasis. 1990;10(1):49–64. [PubMed] [Google Scholar]
  48. Satoh T., Nakafuku M., Kaziro Y. Function of Ras as a molecular switch in signal transduction. J Biol Chem. 1992 Dec 5;267(34):24149–24152. [PubMed] [Google Scholar]
  49. Sekine A., Fujiwara M., Narumiya S. Asparagine residue in the rho gene product is the modification site for botulinum ADP-ribosyltransferase. J Biol Chem. 1989 May 25;264(15):8602–8605. [PubMed] [Google Scholar]
  50. Self A. J., Paterson H. F., Hall A. Different structural organization of Ras and Rho effector domains. Oncogene. 1993 Mar;8(3):655–661. [PubMed] [Google Scholar]
  51. Small J. V. The actin cytoskeleton. Electron Microsc Rev. 1988;1(1):155–174. doi: 10.1016/s0892-0354(98)90010-7. [DOI] [PubMed] [Google Scholar]
  52. Smith D. B., Johnson K. S. Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase. Gene. 1988 Jul 15;67(1):31–40. doi: 10.1016/0378-1119(88)90005-4. [DOI] [PubMed] [Google Scholar]
  53. Sosnowski R. G., Feldman S., Feramisco J. R. Interference with endogenous ras function inhibits cellular responses to wounding. J Cell Biol. 1993 Apr;121(1):113–119. doi: 10.1083/jcb.121.1.113. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Stasia M. J., Jouan A., Bourmeyster N., Boquet P., Vignais P. V. ADP-ribosylation of a small size GTP-binding protein in bovine neutrophils by the C3 exoenzyme of Clostridium botulinum and effect on the cell motility. Biochem Biophys Res Commun. 1991 Oct 31;180(2):615–622. doi: 10.1016/s0006-291x(05)81110-6. [DOI] [PubMed] [Google Scholar]
  55. Stoker M. Effect of scatter factor on motility of epithelial cells and fibroblasts. J Cell Physiol. 1989 Jun;139(3):565–569. doi: 10.1002/jcp.1041390316. [DOI] [PubMed] [Google Scholar]
  56. Stoker M., Gherardi E., Perryman M., Gray J. Scatter factor is a fibroblast-derived modulator of epithelial cell mobility. Nature. 1987 May 21;327(6119):239–242. doi: 10.1038/327239a0. [DOI] [PubMed] [Google Scholar]
  57. Stoker M., Perryman M. An epithelial scatter factor released by embryo fibroblasts. J Cell Sci. 1985 Aug;77:209–223. doi: 10.1242/jcs.77.1.209. [DOI] [PubMed] [Google Scholar]
  58. Takaishi K., Kikuchi A., Kuroda S., Kotani K., Sasaki T., Takai Y. Involvement of rho p21 and its inhibitory GDP/GTP exchange protein (rho GDI) in cell motility. Mol Cell Biol. 1993 Jan;13(1):72–79. doi: 10.1128/mcb.13.1.72. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Takaishi K., Sasaki T., Kato M., Yamochi W., Kuroda S., Nakamura T., Takeichi M., Takai Y. Involvement of Rho p21 small GTP-binding protein and its regulator in the HGF-induced cell motility. Oncogene. 1994 Jan;9(1):273–279. [PubMed] [Google Scholar]
  60. Tominaga T., Sugie K., Hirata M., Morii N., Fukata J., Uchida A., Imura H., Narumiya S. Inhibition of PMA-induced, LFA-1-dependent lymphocyte aggregation by ADP ribosylation of the small molecular weight GTP binding protein, rho. J Cell Biol. 1993 Mar;120(6):1529–1537. doi: 10.1083/jcb.120.6.1529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. Trahey M., Milley R. J., Cole G. E., Innis M., Paterson H., Marshall C. J., Hall A., McCormick F. Biochemical and biological properties of the human N-ras p21 protein. Mol Cell Biol. 1987 Jan;7(1):541–544. doi: 10.1128/mcb.7.1.541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  62. Weidner K. M., Arakaki N., Hartmann G., Vandekerckhove J., Weingart S., Rieder H., Fonatsch C., Tsubouchi H., Hishida T., Daikuhara Y. Evidence for the identity of human scatter factor and human hepatocyte growth factor. Proc Natl Acad Sci U S A. 1991 Aug 15;88(16):7001–7005. doi: 10.1073/pnas.88.16.7001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  63. Weidner K. M., Behrens J., Vandekerckhove J., Birchmeier W. Scatter factor: molecular characteristics and effect on the invasiveness of epithelial cells. J Cell Biol. 1990 Nov;111(5 Pt 1):2097–2108. doi: 10.1083/jcb.111.5.2097. [DOI] [PMC free article] [PubMed] [Google Scholar]

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