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Microtubule growth activates Rac1 to promote lamellipodial protrusion in fibroblasts

Nature Cell Biology volume 1, pages 45–50 (1999)Cite this article

Abstract

Microtubules are involved in actin-based protrusion at the leading-edge lamellipodia of migrating fibroblasts. Here we show that the growth of microtubules induced in fibroblasts by removal of the microtubule destabilizer nocodazole activates Rac1 GTPase, leading to the polymerization of actin in lamellipodial protrusions. Lamellipodial protrusions are also activated by the rapid growth of a disorganized array of very short microtubules induced by the microtubule-stabilizing drug taxol. Thus, neither microtubule shortening nor long-range microtubule-based intracellular transport is required for activating protrusion. We suggest that the growth phase of microtubule dynamic instability at leading-edge lamellipodia locally activates Rac1 to drive actin polymerization and lamellipodial protrusion required for cell migration.

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Figure 1: Microtubule and cellular dynamics in cells after washout of 10 µM nocodazole in an NIH3T3 fibroblast.
Figure 2: Quantification of lamellipodial protrusion, cell spreading and microtubule organization during recovery from nocodazole treatment in fibroblasts.
Figure 3: Microtubule and cellular dynamics in an NIH3T3 cell during exchange from medium containing 10 µM nocodazole into medium containing 10 µM taxol.
Figure 4: F-actin dynamics during nocodazole washout in a Swiss 3T3 cell.
Figure 5: Nocodazole washout in a Swiss 3T3 cell injected with Rac1N17.
Figure 6: Microtubule growth activates Rac1.

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References

  1. Heath, J. P. & Holifield, B. F. Cell locomotion: new research tests old ideas on membrane and cytoskeletal flow. Cell Motil. Cytoskeleton 18, 245–257 (1991).

    Article  CAS  Google Scholar 

  2. Mitchison, T. J. & Cramer, L. P. Actin-based cell motility and cell locomotion. Cell 84, 371–379 (1996).

    Article  CAS  Google Scholar 

  3. Desai, A. & Mitchison, T. J. Microtubule polymerization dynamics. Annu. Rev. Cell Dev. Biol. 13, 83–117 (1997).

    Article  CAS  Google Scholar 

  4. Vasiliev, J. M. et al. Effect of colcemid on the locomotory behaviour of fibroblasts . J. Embryol. Exp. Morph. 24, 625– 690 (1970).

    CAS  Google Scholar 

  5. Bershadsky, A. D., Vaisberg, E. A. & Vasiliev, J. M. Pseudopodial activity at the active edge of migrating fibroblast is decreased after drug-induced microtubule depolymerization. Cell Motil. Cytoskeleton 19, 152–158 (1991).

    Article  CAS  Google Scholar 

  6. Vasiliev, J. M. Polarization of pseudopodial activities: cytoskeletal mechanisms. J. Cell Sci. 98, 1–4 ( 1991).

    PubMed Central  Google Scholar 

  7. Rinnerthaler, G., Geiger, B. & Small, J. Contact formation during fibroblast locomotion: involvement of membrane ruffles and microtubules. J. Cell Biol. 106, 747–760 (1988).

    Article  CAS  Google Scholar 

  8. Rodionov, V. I. et al. Microtubule-dependent control of cell shape and pseudopodial activity is inhibited by the antibody to kinesin motor domain. J. Cell Biol. 123, 1811–1820 (1993).

    Article  CAS  Google Scholar 

  9. Bretscher, M. S. Getting membrane flow and the cytoskeleton to cooperate. Cell 87, 601–606 (1996).

    Article  CAS  Google Scholar 

  10. Singer, S. J. & Kupfer, A. The directed migration of eukaryotic cells. Annu. Rev. Cell Biol. 2, 337– 365 (1986).

    Article  CAS  Google Scholar 

  11. Liao, G., Nagasaki, T. & Gundersen, G. G. Low concentrations of nocodazole interfere with fibroblast locomotion without significantly affecting microtubule level: implications for the role of dynamic microtubules in cell locomotion. J. Cell Sci. 108, 3473–3483 ( 1995).

    CAS  Google Scholar 

  12. Waterman-Storer, C. M. & Salmon, E. D. Actomyosin-based retrograde flow of microtubules in the lamella of migrating epithelial cells influences microtubule dynamic instability and turnover and is associated with microtubule breakage and treadmilling. J. Cell Biol. 139, 417 –434 (1997).

    Article  CAS  Google Scholar 

  13. Bloom, G. S. & Goldstein, L. S. Cruising along microtubule highways: how membranes move through the secretory pathway. J. Cell Biol. 140, 1277–1280 (1998).

    Article  CAS  Google Scholar 

  14. Schiff, P. B. & Horwitz, S. B. Taxol stabilizes microtubules in mouse fibroblast cells. Proc. Natl Acad. Sci. USA 77, 1561–1565 (1980).

    Article  CAS  Google Scholar 

  15. 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 70, 401–410 (1992).

    Article  CAS  Google Scholar 

  16. Machesky, L. M. & Hall, A. Role of actin polymerization and adhesion to extracellular matrix in Rac- and Rho-induced cytoskeletal reorganization. J. Cell Biol. 138, 913– 926 (1997).

    Article  CAS  Google Scholar 

  17. Van Aelst, L. & D’Souza-Schorey, C. Rho GTPases and signaling networks. Genes Dev. 11, 2295– 2322 (1997).

    Article  CAS  Google Scholar 

  18. Nobes, C. D., Hawkins, P., Stephens, L. & Hall, A. Activation of the small GTP-binding proteins rho and rac by growth factor receptors. J. Cell Sci. 108, 225–233 (1995).

    CAS  Google Scholar 

  19. Nobes, C. D. & Hall, A. Rho, rac, and cdc42 GTPases regulate the assembly of multimolecular focal complexes associated with actin stress fibers, lamellipodia, and filopodia. Cell 81, 53–62 (1995).

    Article  CAS  Google Scholar 

  20. Bagrodia, S., Taylor, S. J., Jordon, K.A., Van Aelst, L. & Cerione, R. A. A novel regulator of p21-activated kinases. J. Biol. Chem 273, 23633– 23636 (1998).

    Article  CAS  Google Scholar 

  21. Burbelo, P. D., Drechsel, D. & Hall, A. A conserved binding motif defines numerous candidate target proteins for both Cdc42 and Rac GTPases. J. Biol. Chem. 270, 29071–29074 ( 1995).

    Article  CAS  Google Scholar 

  22. Best, A., Ahmed, S., Kozma, R. & Lim, L. The Ras-related GTPase Rac1 binds tubulin. J. Biol. Chem 271, 3756 –3762 (1996).

    Article  CAS  Google Scholar 

  23. Ren, Y., Li, R., Zheng, Y. & Busch, H. Cloning and characterization of GEF-H1, a microtubule-associated guanine nucleotide exchange factor for Rac and Rho GTPases. J. Biol. Chem. 273, 34954–34960 (1998).

    Article  CAS  Google Scholar 

  24. Fernandez, J. A. et al. Phosphorylation- and actvitation-independent association of the tyrosine kinase Syk and the tyrosine kinase substrates Cbl and Vav with tubulin in B-cells. J. Biol. Chem. 274, 1401–1406 (1999).

    Article  CAS  Google Scholar 

  25. Glaven, J. A., Whitehead, I., Bagrodia, S., Kay, R. & Cerione, R. A. The Dbl-related protein, Lfc, localizes to microtubules and mediates the activation of Rac signaling pathways in cells. J. Biol. Chem 274, 2279-2285 (1999).

    Article  Google Scholar 

  26. Nagata, K. et al. The MAP kinase kinase kinase MLK2 co-localizes with activated JNK along microtubules and associates with kinesin superfamily motor KIF3 . EMBO J. 17, 149–158 (1998).

    Article  CAS  Google Scholar 

  27. Perez, F., Diamantopoulos, G. S., Stalder, R. & Kreis, T. E. Clip-170 highlights growing microtubule ends in vivo. Cell 96, 517–552 ( 1999).

    Article  CAS  Google Scholar 

  28. Gauthier-Rouviere, C. et al. RhoG GTPase controls a pathway that independently activates Rac1 and Cdc42 Hs. Mol. Biol. Cell 9, 1379 –1394 (1998).

    Article  CAS  Google Scholar 

  29. Enomoto, T. Microtubule disruption indcues the formation of actin stress fibers and focal adhesions in cultured cells: possible involvement of the Rho signal cascade . Cell Struct. Funct. 21, 317– 326 (1996).

    Article  CAS  Google Scholar 

  30. Zhang, Q., Magnusson, M. K. & Mosher, D. F. Lysophosphatidic acid and microtubule-destabilizing agents stimulate fibronectin matrix assembly through Rho-dependent actin stress fiber formation and cell contraction,. Mol. Biol. Cell 8, 1415–1425 (1997).

    Article  CAS  Google Scholar 

  31. Liu, B., Chrzanowska-Wodnicka, M. & Burridge, K. Microtubule depolymerization induces stress fibers, focal adhesions, and DNA synthesis via the GTP-binding protein Rho . Cell Adhesion Commun. 5, 249– 255 (1998).

    Article  CAS  Google Scholar 

  32. Ren, X.-D., Kiosses, W. B. & Schwartz, M. A. Regulation of the small GTP-binding protein Rho by cell adhesion and the cytoskeleton. EMBO J. 18, 578–585 (1999).

    Article  CAS  Google Scholar 

  33. Waterman-Storer, C. M. & Salmon, E. D. Positive feedback interactions between microtubule and actin dynamics during cell motility. Curr. Opin. Cell Biol. 11, 61–67 (1999).

    Article  CAS  Google Scholar 

  34. Waterman-Storer, C. M., Desai, A., Bulinski, J. C. & Salmon, E. D. Fluorescent speckle microscopy, a method to visualize the dynamics of protein assemblies in living cells. Curr. Biol. 8, 1227–1230 (1998).

    Article  CAS  Google Scholar 

  35. Self, A. J. & Hall, A. Purification of recombinant Rho/Rac/G25K from Escherichia coli. Methods Enzymol. 256, 3–10 (1995).

    Article  CAS  Google Scholar 

  36. Salmon, E. D. et al. A high-resolution multimode digital microscope system. Methods Cell Biol. 56, 185–215 (1998).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank S. Earp, C. Der, S. Bagrodia, R. Cerione, B. Kreft, A. Ridley and D. Cheney for their kind contributions of reagents and advice. C.M.W.-S. is a fellow of the Jane Coffin Childs Fund; R.A.W is supported by an NIH grant to the Lineberger Cancer Center. This work was also supported by NIH grants to E.D.S. and K.B.

Correspondence and requests for materials should be addressed to C.M.W.-S.

Supplementary information is available on Nature Cell Biology’s World-Wide Web site (http://www.nature.com/ncb/webfocus/index.html).

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Authors and Affiliations

  1. Department of Biology, University of North Carolina, Chapel Hill, 27599, North Carolina, USA

    Clare M. Waterman-Storer & E.D. Salmon

  2. Department of Cell Biology/Anatomy and Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, 27599, North Carolina, USA

    Rebecca A. Worthylake, Betty P. Liu & Keith Burridge

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  1. Clare M. Waterman-Storer
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Correspondence to Clare M. Waterman-Storer.

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Waterman-Storer, C., Worthylake, R., Liu, B. et al. Microtubule growth activates Rac1 to promote lamellipodial protrusion in fibroblasts. Nat Cell Biol 1, 45–50 (1999). https://doi.org/10.1038/9018

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