Induction of epithelial tubules by growth factor HGF depends on the STAT pathway

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Abstract

Hepatocyte growth factor (HGF) induces a three-phase response leading to the formation of branched tubular structures in epithelial cells1,2. The HGF receptor tyrosine kinase works through a Src homology (SH2) docking site that can activate several signalling pathways3. The first phase of the response (scattering), which results from cytoskeletal reorganization, loss of intercellular junctions and cell migration4, is dependent on phosphatidylinositol-3-OH kinase and Rac activation5,6. The second phase (growth) requires stimulation of the Ras–MAP kinase cascade7. Here we show that the third phase (tubulogenesis) is dependent on the STAT pathway. HGF stimulates recruitment of Stat-3 to the receptor, tyrosine phosphorylation, nuclear translocation and binding to the specific promoter element SIE. Electroporation of a tyrosine-phosphorylated peptide, which interferes with both the association of STAT to the receptor and STAT dimerization, inhibits tubule formation in vitro without affecting either HGF-induced ‘scattering’ or growth. The same result is obtained using a specific ‘decoy’ oligonucleotide that prevents STAT from binding to DNA and affecting the expression of genes involved in cell-cycle regulation (c-fos and waf-1). Activation of signal transducers that directly control transcription is therefore required for morphogenesis.

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Figure 1: HGF induces STAT phosphorylation and translocation into the nucleus.
Figure 2: Stat-3 associates with the HGF receptor.
Figure 3: The SIE elements binds STAT and modulates the expression of HGF-induced genes.
Figure 4: Selective inhibition of branching morphogenesis by a STAT phosphopeptide.

References

  1. 1

    Medico, E. et al. The tyrosine kinase receptors Ron and Sea control “scattering” and morphogenesis of liver progenitor cells in vitro. Mol. Biol. Cell 7, 495–504 (1996).

  2. 2

    Sachs, M. et al. Mitogenic and morphogenic activity of epithelial receptor tyrosine kinases. J. Cell Biol. 133, 1095–1107 (1996).

  3. 3

    Ponzetto, C. et al. Amultifunctional docking site mediates signalling and transformation by the Hepatocyte Growth Factor/Scatter Factor receptor family. Cell 77, 261–271 (1994).

  4. 4

    Stocker, M., Gherardi, E., Perryman, M. & Gray, J. Scatter factor is a fibroblast-derived modulator of epithelial cell motility. Nature 327, 239–242 (1987).

  5. 5

    Ridley, A. J., Comoglio, P. M. & Hall, A. Regulation of Scatter Factor/Hepatocyte Growth Factor responses by Ras, Rac, and Rho in MDCK cells. J. Cell Biol. 15, 1110–1122 (1995).

  6. 6

    Royal, I. & Park, M. Hepatocyte Growth Factor-induced scatter of Madin–Darby canine kidney cells requires phosphatidylinositol 3-kinase. J. Biol. Chem. 270, 27780–27787 (1995).

  7. 7

    Ponzetto, C. et al. Specific uncoupling of GRB2 from the Met receptor. J. Biol. Chem. 271, 14119–14123 (1996).

  8. 8

    Pawson, T. Protein modules and signalling networks. Nature 373, 573–580 (1995).

  9. 9

    Pelicci, G. et al. The motogenic and mitogenic responses to HGF are amplified by the Shc adaptor protein. Oncogene 10, 1631–1638 (1995).

  10. 10

    Weidner, K. M. et al. Interaction between Gab1 and the c-Met receptor tyrosine kinase is responsible for epithelial morphogenesis. Nature 384, 173–176 (1996).

  11. 11

    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. 266, 22087–22090 (1991).

  12. 12

    Ponzetto, C. et al. Anovel recognition motif for Phosphatidylinositol 3-kinase binding mediates its association with the Hepatocyte Growth Factor/Scatter Factor receptor. Mol. Cell Biol. 13, 4600–4608 (1993).

  13. 13

    Graziani, A., Gramaglia, D., dalla Zonca, P. & Comoglio, P. M. Hepatocyte Growth Factor/Scatter Factor stimulates the Ras-guanine nucleotide exchanger. J. Biol. Chem. 268, 9165–9168 (1993).

  14. 14

    Schindler, C. & Darnell, J. E. J Transcriptional responses to polypeptide ligands: the JAK-STAT pathway. Annu. Rev. Biochem. 64, 621–651 (1995).

  15. 15

    Ihle, J. N. STATs: signal transducers and activators of transcription. Cell 84, 221–334 (1996).

  16. 16

    Leaman, D. W. et al. Roles of JAKs in activation of STATs and stimulation of c-fos gene expression by Epidermal Growth Factor. Mol. Cell. Biol. 16, 369–375 (1996).

  17. 17

    Silvennoinen, O., Ihle, J. N., Schlessinger, J. & Levy, D. E. Interferon-induced nuclear signalling by Jak protein tyrosine kinases. Nature 366, 583–585 (1993).

  18. 18

    Wagner, B. J., Hayes, T. E., Hoban, C. J. & Cochran, B. H. The SIF binding element confers sis/PDGF inductibility onto the c-fos promoter. EMBO J. 9, 4477–4484 (1990).

  19. 19

    Durbin, J. E., Hackenmiller, R., Simon, M. C. & Levy, D. E. Targeted disruption of the mouse Stat1 gene results in compromised innate immunity to viral disease. Cell 84, 443–450 (1996).

  20. 20

    Liu, X. et al. Stat5a is mandatory for adult mammary gland development and lactogenesis. Genes Dev. 11, 179–186 (1967).

  21. 21

    Takeda, K. et al. Targeted disruption of the mouse Stat3 gene leads to early embryonic lethality. Proc. Natl Acad. Sci. USA 94, 3801–3804 (1997).

  22. 22

    Chin, Y. E. et al. Cell growth arrest and induction of cyclin-dependent kinase inhibitor p21WAF1/CIP1 mediated by STAT1. Science 272, 719–722 (1996).

  23. 23

    Boccaccio, C., Gaudino, G., Gambarotta, G., Galimi, F. & Comoglio, P. M. Hepatocyte Growth factor (HGF) receptor expression is inducible and is part of the delayed-early response to HGF. J. Biol. Chem. 269, 12846–12851 (1994).

  24. 24

    Lucibello, F. C., Lowag, C., Neuberg, M. & Muller, R. Trans-repression of the mouse c-fos promoter: a novel mechanism of Fos-mediated trans-regulation. Cell 59, 999–1007 (1989).

  25. 25

    Giordano, S., Ponzetto, C., Di Renzo, M. F., Cooper, C. S. & Comoglio, P. M. Tyrosine kinase receptor indistinguishable from the c-Met protein. Nature 339, 155–156 (1989).

  26. 26

    Gilman, M. Z., Wilson, R. N. & Weinberg, R. A. Multiple protein binding sites in the 5′-flanking region regulate c-fos expression. Mol. Cell. Biol. 6, 4305–4316 (1986).

  27. 27

    El-Deiry, W. S. et al. WAF-1, a potential mediator of p53 tumor suppression. Cell 75, 817–825 (1993).

  28. 28

    Gambarotta, G. et al. Ets up-regulates MET transcription. Oncogene 13, 1911–1917 (1996).

  29. 29

    Raptis, L. H., Liu, S. K. W., Firth, K. L., Stiles, C. D. & Alberta, J. A. Electroporation of peptides into adherent cells in situ. Biotechniques 19, 104–114 (1995).

  30. 30

    Comoglio, P. M. et al. Detection of phosphotyrosine containing proteins in the detergent insoluble fraction of RSV-transformed fibroblasts by azobenzene phosphonate antibodies. EMBO J. 3, 483–489 (1984).

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Acknowledgements

We thank P. Giordano (Pharmacia & Upjohn) for peptide synthesis; C. Ponzetto for discussions; A. Cignetto for secretarial help; and E. Wright for help with the manuscript. This work was supported by an AIRC grant to P.M.C. M.A. is recipient of a FIRC Fellowship.

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Correspondence to Carla Boccaccio.

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