Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Paper
  • Published:

VEGF activates protein kinase C-dependent, but Ras-independent Raf-MEK-MAP kinase pathway for DNA synthesis in primary endothelial cells

Oncogene volume 18, pages 2221–2230 (1999)Cite this article

Abstract

KDR/Flk-1 tyrosine kinase, one of the two VEGF receptors induces mitogenesis and differentiation of vascular endothelial cells. We have previously reported that a major target molecule of KDR/Flk-1 kinase is PLC-γ, and that VEGF induces activation of MAP kinase, mainly mediated by protein kinase C (PKC) in the NIH3T3 cells overexpressing KDR/Flk-1 (Takahashi and Shibuya, 1997). However, the signal transduction initiated from VEGF in endothelial cells remains to be elucidated. In primary sinusoidal endothelial cells which showed strictly VEGF-dependent growth, we found that VEGF stimulated the activation of Raf-1-MEK-MAP kinase cascade. To our surprise, an important regulator, Ras was not efficiently activated to a significant level in response to VEGF. Consistent with this, dominant-negative Ras did not block the VEGF-induced phosphorylation of MAP kinase. On the other hand, PKC-specific inhibitors severely reduced VEGF-dependent phosphorylation of MEK, activation of MAP kinase and subsequent DNA synthesis. A potent PI3 kinase inhibitor, Wortmannin, could not inhibit either of them. These results suggest that in primary endothelial cells, VEGF-induced activation of Raf-MEK-MAP kinase and DNA synthesis are mainly mediated by PKC-dependent pathway, much more than by Ras-dependent or PI3 kinase-dependent pathway.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7

Similar content being viewed by others

References

  • Abedi H and Zachary I. . 1997 J. Biol. Chem. 272: 15442–15451.

  • Alessi DR, Cuenda A, Cohen P, Dudley DT and Saltiel AR. . 1995 J. Biol. Chem. 270: 27489–27494.

  • Baccarini M, Sabatini DM, App H, Rapp UR and Stanley ER. . 1990 EMBO J. 9: 3649–3657.

  • Barleon B, Sozzani S, Zhou D, Weich HA, Mantovani A and Marmé D. . 1996 Blood 87: 3336–3343.

  • Cai H, Smola U, Wixler V, Eisenmann-Tappe I, Diaz-Meco MT, Moscat J, Rapp U and Cooper GM. . 1997 Mol. Cell. Biol. 17: 732–741.

  • Clauss HSM, Ryan J, Schmidt A-M, Tijburg P, Borden L, Connolly D, Stern D and Kao J. . 1993 Blood 81: 2767–2773.

  • Clauss M, Weich H, Breier G, Knies U, Röck W, Waltenberger J and Risau W. . 1996 J. Biol. Chem. 271: 17629–17634.

  • Cohen T, Gitay-Goren H, Neufeld G and Levi B-Z. . 1992 Growth Fact. 7: 131–138.

  • Conningham SA, Waxham MN, Arrate PM and Brock TA. . 1995 J. Biol. Chem. 270: 20254–20257.

  • Connolly DT, Heuvelman DM, Nelson R, Olander JV, Eppley BL, Delfino JJ, Siegel NR, Leimgruber RM and Feder J. . 1989 J. Clin. Invest. 84: 1470–1478.

  • Daviet I, Herbert JM and Maffrand JP. . 1989 Biochem. Biophys. Res. Commun. 158: 584–589.

  • De Vries C, Escobedo JA, Ueno H, Houck K, Ferrara N and Williams LT. . 1992 Science 255: 989–991.

  • Eichmann A, Marcella C, Bréant C and Le Douarin NM. . 1993 Mechanisms Dev. 42: 33–48.

  • Ferrara N and Henzel WJ. . 1989 Biochem. Biophys. Res. Commun. 161: 851–858.

  • Ferrara N, Houck KA, Jakeman LB, Winer J and Leung DW. . 1991 J. Cell. Biochem. 47: 211–218.

  • Folkman J. . 1990 J. Natl. Cancer Inst. 82: 4–6.

  • Fong G-H, Rossant J, Gertsenstein M and Breitman ML. . 1995 Nature 376: 66–70.

  • Gotoh N, Tojo A, Murota K, Hashimoto Y, Hattori S, Nakamura S, Takenawa T, Yazaki Y and Shibuya M. . 1994 Proc. Natl. Acad. Sci. USA 91: 167–171.

  • Gotoh Y, Nishida E, Yamashita T, Hoshi M, Kawakami M and Sakai H. . 1990 Eur. J. Biochem. 193: 661–669.

  • Guo D, Jia Q, Song H-Y, Warren RS and Donner DB. . 1995 J. Biol. Chem. 270: 6729–6733.

  • Inoguchi T, Battan R, Handler E, Sportsman JR, Health W and King GL. . 1992 Proc. Natl. Acad. Sci. USA 89: 11059–11063.

  • Izumi T, Tamemoto H, Nagao M, Kadowaki T, Takaku F and Kasuga M. . 1991 J. Biol. Chem. 266: 7933–7939.

  • Jakeman LB, Winer J, Bennett GL, Altar CA and Ferrara N. . 1993 J. Clin. Invest. 89: 244–253.

  • Jung V, Wei W, Ballester R, Camonis J, Mi S, Van Aelst L, Wigler M and Broek D. . 1994 Mol. Cell. Biol. 14: 3707–3718.

  • Kaipainen A, Korhonen J, Pajusola K, Aprelikova O, Persico MG, Terman BI and Alitalo K. . 1993 J. Exp. Med. l78: 2077–2088.

  • Katagiri H, Asano T, Inukai K, Ogihara T, Ishihara H, Shibasaki Y, Murata T, Terasaki J, Kikuchi M, Yazaki Y and Oka Y. . 1997 Am. J. Physiol. 272: E326–331.

  • Katoh O, Tauchi H, Kawaishi K, Kimura A and Satow Y. . 1995 Cancer Res. 55: 5687–5692.

  • Keck PJ, Hauser SD, Krivi G, Sanzo K, Warren T, Feder J and Connolly DT. . 1989 Science 246: 1309–1312.

  • Kolch W, Heldecker G, Kochs G, Hummel R, Vahidi H, Mischak H, Finkenzeller G, Marmé D and Rapp UR. . 1993 Nature 364: 249–252.

  • Kroll J and Waltenberger J. . 1997 J. Biol. Chem. 272: 33521–33527.

  • Leung DW, Cachianes G, Kuang W-J, Goeddel DV and Ferrara N. . 1989 Science 246: 1306–1309.

  • Li Y-S, Shyy JY-J, Li S, Lee J, Su B, Karin M and Chien S. . 1996 Mol. Cell. Biol. 16: 5947–5954.

  • Martiny-Baron G, Kazanietz MG, Mischak H, Blumberg PM, Kochs G, Hug H, Marmé D and Schächtele C. . 1993 J. Biol. Chem. 268: 9194–9197.

  • Matthews W, Jordan CT, Gavin M, Jenkins NA, Copeland NG and Lemischka IR. . 1991 Proc. Natl. Acad. Sci. USA 88: 9026–9030.

  • Millauer B, Wizigmann-Voos S, Schnürch H, Martinez R, Moller NPH, Risau W and Ullrich A. . 1993 Cells 72: 835–846.

  • Miyake S, Makimura M, Kanegae Y, Harada S, Sato Y, Takamori K, Tokuda C and Saito I. . 1996 Proc. Natl. Acad. Sci. USA 93: 1320–1324.

  • Mochly-Rosen D. . 1995 Science 268: 247–251.

  • Montesano R and Orci L. . 1985 Cell 42: 469–477.

  • Morris PB, Hida T, Blackshear PJ, Klintworth GK and Swain JL. . 1988 Am. J. Physiol. 254: C318–C322.

  • Muroya K, Hattori S and Nakamura S. . 1992 Oncogene 7: 277–281.

  • Mustonen T and Alitalo K. . 1995 J. Cell. Biol. 12: 895–898.

  • Nishizuka Y. . 1992 Science 258: 607–614.

  • Pepper MS, Ferrara N, Orci L and Montesano R. . 1991 Biochem. Biophys. Res. Commun. 181: 902–906.

  • Qiu M-S and Green SH. . 1991 Neuron 7: 937–946.

  • Quinn TP, Peter KG, De Vries C, Ferrara N and Williams LT. . 1993 Proc. Natl. Acad. Sci. USA 90: 7533–7537.

  • Satoh T, Endo M, Nakafuku M, Nakamura S and Kaziro Y. . 1990a Proc. Natl. Acad. Sci. USA 87: 5993–5997.

  • Satoh T, Endo M, Nakafuku M, Akiyama T, Yamamoto T and Kaziro Y. . 1990b Proc. Natl. Acad. Sci. USA 87: 7926–7929.

  • Sawano A, Takahashi T, Yamaguchi S and Shibuya M. . 1997 Biochem. Biophys. Res. Commun. 238: 487–491.

  • Seetharam L, Gotoh N, Maru Y, Neufeld G, Yamaguchi S and Shibuya M. . 1995 Oncogene 10: 135–147.

  • Senger DR, Galli SJ, Dvorak AM, Perruzzi CA, Harvey VS and Dvorak HF. . 1983 Science 219: 983–985.

  • Shalaby F, Rossant J, Yamaguchi TP, Gertsenstein M, Wu X-F, Breitman ML and Schuh AC. . 1995 Nature 376: 62–66.

  • Shibuya M. . 1995 Adv. Cancer Res. 67: 281–316.

  • Shibuya M, Yamaguchi S, Yamane A, Ikeda T, Tojo A, Matushime H and Sato M. . 1990 Oncogene 5: 519–524.

  • Sözeri O, Vollmer K, Liyanage M, Frith D, Kour G, Mark III GE and Stabel S. . 1992 Oncogene 7: 2259–2267.

  • Takahashi T, Kuwahara Y, Okuda M, Ueno H, Takeshita A and Yokoyama M. . 1997 J. Biol. Chem. 272: 16018–16022.

  • Takahashi T and Shibuya M. . 1997 Oncogene 14: 2079–2089.

  • Terman BI, Dougher-Vermazen, Carrion ME, Dimitrov D, Armellino DC, Gospodarowicz D and Böhlen P. . 1992 Biochem. Biophys. Res. Commun. 187: 1579–1586.

  • Toullec D, Pianetti P, Coste H, Bellevengue P, Grand-Perret T, Ajakane M, Baudet V, Boissin P, Boursier E, Loriolle F, Duhamel L, Charon D and Kirilovsky J. . 1991 J. Biol. Chem. 266: 15771–15781.

  • Ueda Y, Hirai S, Osada S, Suzuki A, Mizuno K and Ohno S. . 1996 J. Biol. Chem. 271: 23512–23519.

  • Unemori EN, Ferrara N, Bauer EA and Amento EP. . 1992 J. Cell. Phys. 153: 557–562.

  • Waltenberger J, Claesson-Welsh L, Siegbahn A, Shibuya M and Heldin C-H. . 1994 J. Biol. Chem. 269: 26988–26995.

  • Wolf BA, Williamson JR, Easom RA, Chang K, Sherman WR and Turk J. . 1991 J. Clin. Invest. 87: 31–38.

  • Xia P, Aiello LP, Ishii H, Jiang ZY, Park DJ, Robinson GS, Takagi H, Newsome WP, Jirousek MR and King GL. . 1996 J. Clin. Invest. 98: 2018–2026.

  • Yamane A, Seetharam L, Yamaguchi S, Gotoh N, Takahashi T, Neufeld G and Shibuya M. . 1994 Oncogene 9: 2683–2690.

Download references

Acknowledgements

We would like to thank Drs Eisuke Nishida, Hitoshi Ito and Hiroshi Koide for supplying Raf kinase assay system; Drs Yoshitomo Oka and Hideki Katagiri for the adenoviral vector expressing Ki-Ras17N; Dr Yasuhisa Fukui for monoclonal antibody against p85 subunit of PI3 kinase; Dr Tadaomi Takenawa for anti-Grb2 serum. We are also grateful to Mrs S Yamaguchi and A Sawano for excellent technical assistance. This work was supported by Grant-in-Aid for Special Project Research on Cancer-Bioscience 04253204 from the Ministry of Education and by research grants from the Yakult Bioscience Foundation, the Mitsubishi Foundation and the Foundation for Promotion of Cancer Research in Japan.

Author information

Authors and Affiliations

  1. Department of Genetics, Institute of Medical Science, University of Tokyo Minato-ku, Tokyo, 108-8639, Japan

    Tomoko Takahashi & Masabumi Shibuya

  2. Department of Cardiology, Kyushu University School of Medicine, Fukuoka, 812-0054, Japan

    Hikaru Ueno

Authors
  1. Tomoko Takahashi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hikaru Ueno
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Masabumi Shibuya
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Takahashi, T., Ueno, H. & Shibuya, M. VEGF activates protein kinase C-dependent, but Ras-independent Raf-MEK-MAP kinase pathway for DNA synthesis in primary endothelial cells. Oncogene 18, 2221–2230 (1999). https://doi.org/10.1038/sj.onc.1202527

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.onc.1202527

Keywords

This article is cited by

Search

Advanced search

Quick links