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.

  • Letter
  • Published:

Differentially oriented populations of actin filaments generated in lamellipodia collaborate in pushing and pausing at the cell front

Abstract

Eukaryotic cells advance in phases of protrusion, pause and withdrawal1. Protrusion occurs in lamellipodia, which are composed of diagonal networks of actin filaments, and withdrawal terminates with the formation of actin bundles parallel to the cell edge. Using correlated live-cell imaging and electron microscopy, we have shown that actin filaments in protruding lamellipodia subtend angles from 15–90° to the front, and that transitions from protrusion to pause are associated with a proportional increase in filaments oriented more parallel to the cell edge. Microspike bundles of actin filaments also showed a wide angular distribution and correspondingly variable bilateral polymerization rates along the cell front. We propose that the angular shift of filaments in lamellipodia serves in adapting to slower protrusion rates while maintaining the filament densities required for structural support; further, we suggest that single filaments and microspike bundles contribute to the construction of the lamella behind and to the formation of the cell edge when protrusion ceases. Our findings provide an explanation for the variable turnover dynamics of actin filaments in lamellipodia observed by fluorescence speckle microscopy2 and are inconsistent with a current model of lamellipodia structure that features actin filaments branching at 70° in a dendritic array3.

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

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

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

Figure 1: Arrest of a steadily protruding lamellipodium and preservation of the actin gradient.
Figure 2: Actin filaments are variably orientated in protruding lamellipodia.
Figure 3: Actin filaments reorganize during the transition from protrusion to retraction.
Figure 4: Slowing and pause is associated with an increase in the number of filaments at shallow angles to the lamellipodium front.
Figure 5: Filament reorganization during protrusion, pause and retraction.

Similar content being viewed by others

References

  1. Abercrombie, M., Heaysman, J.E. & Pegrum, S. M. The locomotion of fibroblasts in culture. II. 'Ruffling'. Exp. Cell Res. 60, 437–444 (1970).

    Article  CAS  Google Scholar 

  2. Ponti, A., Machacek, M. S., Gupton, L., Waterman-Storer, C. M. & Danuser, G. Two distinct actin networks drive the protrusion of migrating cells. Science 305, 1782–1786 (2004).

    Article  CAS  Google Scholar 

  3. Svitkina, T. M. & Borisy, G. G. Arp2/3 complex and actin depolymerizing factor/cofilin in dendritic organization and treadmilling of actin filament array in lamellipodia. J. Cell Biol. 145, 1009–1026 (1999).

    Article  CAS  Google Scholar 

  4. Small, J. V., Stradal, T., Vignal, E. & Rottner, K. The lamellipodium: where motility begins. Trends Cell Biol. 12, 112–20 (2002).

    Article  CAS  Google Scholar 

  5. Small, J. V., Isenberg, G. & Celis, J. E. Polarity of actin at the leading edge of cultured cells. Nature 272, 638–639 (1978).

    Article  CAS  Google Scholar 

  6. Small, J. V., Herzog, M. & Anderson, K. Actin filament organization in the fish keratocyte lamellipodium. J. Cell Biol. 129, 1275–1286 (1995).

    Article  CAS  Google Scholar 

  7. Pollard, T. D. & Borisy, G. Cellular motility driven by assembly and disassembly of actin filaments. Cell 112, 453–465 (2003).

    Article  CAS  Google Scholar 

  8. Pollard, T. D. Regulation of actin filament assembly by arp2/3 complex and formins. Annu. Rev. Biophys. Biomol. Struct. 36, 451–477 (2007).

    Article  CAS  Google Scholar 

  9. Heath, J. P. & Holifield, B. F. On the mechanisms of cortical actin flow and its role in cytoskeletal organisation of fibroblasts. Symp. Soc. Exp. Biol. 47, 35–56 (1993).

    CAS  PubMed  Google Scholar 

  10. Rottner, K., Behrendt, B., Small, J. V. & Wehland, J. VASP dynamics during lamellipodia protrusion. Nature Cell Biol. 1, 321–322 (1999).

    Article  CAS  Google Scholar 

  11. Condeelis, J. & Segall, J. E. Intravital imaging of cell movement in tumours. Nature Rev. Cancer 3, 921–930 (2003).

    Article  CAS  Google Scholar 

  12. Small, J. V. & Resch, G. P. The comings and goings of actin: coupling protrusion and retraction in cell motility. Curr. Opin. Cell Biol. 17, 517–523 (2005).

    Article  CAS  Google Scholar 

  13. Medeiros, N. A., Burnette, D. T. & Forscher, P. Myosin II functions in actin-bundle turnover in neuronal growth cones. Nature Cell Biol. 8, 215–226 (2006).

    Article  CAS  Google Scholar 

  14. Small, J. V. The actin cytoskeleton. Electron Microsc. Rev. 1, 155–174 (1988).

    Article  CAS  Google Scholar 

  15. Abraham, V. C., Krishnamurthi, V., Taylor, D. L. & Lanni, F. The actin-based nanomachine at the leading edge of migrating cells. Biophys. J. 77, 1721–1732 (1999).

    Article  CAS  Google Scholar 

  16. Kozlov, M. M. & Bershadsky, A. D. Processive capping by formin suggests a force-driven mechanism of actin polymerization. J. Cell Biol. 167, 1011–1017 (2004).

    Article  CAS  Google Scholar 

  17. Carlier, M. F. & Pantaloni, D. Control of actin assembly dynamics in cell motility. J. Biol. Chem. 282, 23005–23009 (2007).

    Article  CAS  Google Scholar 

  18. Mogilner, A. On the edge: modeling protrusion. Curr. Opin. Cell Biol. 18, 32–39 (2006).

    Article  CAS  Google Scholar 

  19. Fisher, G. W., Conrad, P. A., DeBiasio, R. L. & Taylor, D. L. Centripetal transport of cytoplasm, actin, and the cell surface in lamellipodia of fibroblasts. Cell Motil. Cytoskeleton 11, 235–247 (1988).

    Article  CAS  Google Scholar 

  20. Vallotton, P., Danuser, G., Bohnet, S., Meister, J. J. & Verkhovsky, A. B. Tracking retrograde flow in keratocytes: news from the front. Mol. Biol. Cell 16, 1223–1231 (2005).

    Article  CAS  Google Scholar 

  21. Hotulainen, P. & Lappalainen, P. Stress fibers are generated by two distinct actin assembly mechanisms in motile cells. J. Cell Biol. 173, 383–394 (2006).

    Article  CAS  Google Scholar 

  22. Blanchoin, L., Pollard, T. D. & Hitchcock-DeGregori, S. E. Inhibition of the Arp2/3 complex-nucleated actin polymerization and branch formation by tropomyosin. Curr. Biol. 11, 1300–1304 (2001).

    Article  CAS  Google Scholar 

  23. Pollard, T. D., Blanchoin, L. & Mullins, R. D. Actin dynamics. J. Cell Sci. 114, 3–4 (2001).

    CAS  PubMed  Google Scholar 

  24. Amann, K. J. & Pollard, T. D. Direct real-time observation of actin filament branching mediated by Arp2/3 complex using total internal reflection fluorescence microscopy. Proc. Natl Acad. Sci. USA. 98, 15009–15013 (2001).

    Article  CAS  Google Scholar 

  25. Mullins, R.D., Heuser, J. A. & Pollard, T. D. The interaction of Arp2/3 complex with actin: nucleation, high affinity pointed end capping, and formation of branching networks of filaments. Proc. Natl Acad. Sci. USA. 95, 6181–6186 (1998).

    Article  CAS  Google Scholar 

  26. Resch, G. P., Goldie, K. N., Hoenger, A., & Small, J. V. Pure F-actin networks are distorted and branched by steps in the critical-point drying method. J. Struct. Biol. 137, 305–312 (2002).

    Article  CAS  Google Scholar 

  27. Medalia, O., Weber, I., Frangakis, A. S., Nicastro, D., Gerisch, G. & Baumeister, W. Macromolecular architecture in eukaryotic cells visualized by cryoelectron tomography. Science. 298, 1209–1213 (2002).

    Article  CAS  Google Scholar 

  28. Gupton, S.L. et al. Cell migration without a lamellipodium: translation of actin dynamics into cell movement mediated by tropomyosin. J. Cell Biol. 168, 619–631 (2005).

    Article  CAS  Google Scholar 

  29. Giannone, G. et al. Lamellipodial actin mechanically links myosin activity with adhesion-site formation. Cell 128, 561–575 (2007).

    Article  CAS  Google Scholar 

  30. Hebert, B., Costantino, S. & Wiseman, P. W. Spatiotemporal image correlation spectroscopy (STICS) theory, verification, and application to protein velocity mapping in living CHO cells. Biophys. J. 88, 3601–3614 (2005).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors thank the Human Frontier Science Program Organization (HFSPO), The Austrian Science Research Council (FWF) and the Vienna Science Research and Technology Fund (WWTF) as well as the City of Vienna/Zentrum für Innovation und Technologie via the Spot of Excellence grant 'Center of Molecular and Cellular Nanostructure' for financial support. K.R. was supported in part by grants from the Deutsche Forschungsgemeinschaft (SPP1150 and FOR629). We also thank Guenter Resch for the electron microscope facility management and advice with image processing, Tibor Kulcsar and Hannes Tkadletz for graphics and Natalia Andreyeva for helpful comments. The authors thank Roger Tsien, Annette Muller-Taubenberger, Malgorzata Szczodrak, George Patterson, Jennifer Lippincott-Schwarz and Rex Chisholm for probes, and Jeff Segall and Bob van de Water for MTLn3 cells.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to J. Victor Small.

Supplementary information

Supplementary Information

Supplementary figures S1, S2 and S3 (PDF 13184 kb)

Supplementary Information

Supplementary Information Movie 1 (MOV 1466 kb)

Supplementary Information

Supplementary Information Movie 2 (MOV 4009 kb)

Supplementary Information

Supplementary Information Movie 3 (MOV 6633 kb)

Supplementary Information

Supplementary Information Movie 4 (MOV 9398 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Koestler, S., Auinger, S., Vinzenz, M. et al. Differentially oriented populations of actin filaments generated in lamellipodia collaborate in pushing and pausing at the cell front. Nat Cell Biol 10, 306–313 (2008). https://doi.org/10.1038/ncb1692

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ncb1692

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing