Skip to main content
PMC home page
Biophysical Journal logoLink to Biophysical Journal
. 1999 Sep;77(3):1721–1732. doi: 10.1016/S0006-3495(99)77018-9

The actin-based nanomachine at the leading edge of migrating cells.

V C Abraham 1, V Krishnamurthi 1, D L Taylor 1, F Lanni 1
PMCID: PMC1300458  PMID: 10465781

Abstract

Two fundamental parameters of the highly dynamic, ultrathin lamellipodia of migrating fibroblasts have been determined-its thickness in living cells (176 +/- 14 nm), by standing-wave fluorescence microscopy, and its F-actin density (1580 +/- 613 microm of F-actin/microm(3)), via image-based photometry. In combination with data from previous studies, we have computed the density of growing actin filament ends at the lamellipodium margin (241 +/- 100/microm) and the maximum force (1.86 +/- 0.83 nN/microm) and pressure (10.5 +/- 4.8 kPa) obtainable via actin assembly. We have used cell deformability measurements (. J. Cell Sci. 44:187-200;. Proc. Natl. Acad. Sci. USA. 79:5327-5331) and an estimate of the force required to stall the polymerization of a single filament (. Proc. Natl. Acad. Sci. USA. 78:5613-5617;. Biophys. J. 65:316-324) to argue that actin assembly alone could drive lamellipodial extension directly.

Full Text

The Full Text of this article is available as a PDF (553.1 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Abercrombie M., Heaysman J. E., Pegrum S. M. The locomotion of fibroblasts in culture. IV. Electron microscopy of the leading lamella. Exp Cell Res. 1971 Aug;67(2):359–367. doi: 10.1016/0014-4827(71)90420-4. [DOI] [PubMed] [Google Scholar]
  2. Azuma T., Witke W., Stossel T. P., Hartwig J. H., Kwiatkowski D. J. Gelsolin is a downstream effector of rac for fibroblast motility. EMBO J. 1998 Mar 2;17(5):1362–1370. doi: 10.1093/emboj/17.5.1362. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bailey B., Farkas D. L., Taylor D. L., Lanni F. Enhancement of axial resolution in fluorescence microscopy by standing-wave excitation. Nature. 1993 Nov 4;366(6450):44–48. doi: 10.1038/366044a0. [DOI] [PubMed] [Google Scholar]
  4. Cano M. L., Cassimeris L., Joyce M., Zigmond S. H. Characterization of tetramethylrhodaminyl-phalloidin binding to cellular F-actin. Cell Motil Cytoskeleton. 1992;21(2):147–158. doi: 10.1002/cm.970210208. [DOI] [PubMed] [Google Scholar]
  5. Chan A. Y., Raft S., Bailly M., Wyckoff J. B., Segall J. E., Condeelis J. S. EGF stimulates an increase in actin nucleation and filament number at the leading edge of the lamellipod in mammary adenocarcinoma cells. J Cell Sci. 1998 Jan;111(Pt 2):199–211. doi: 10.1242/jcs.111.2.199. [DOI] [PubMed] [Google Scholar]
  6. Condeelis J. Life at the leading edge: the formation of cell protrusions. Annu Rev Cell Biol. 1993;9:411–444. doi: 10.1146/annurev.cb.09.110193.002211. [DOI] [PubMed] [Google Scholar]
  7. Conrad P. A., Nederlof M. A., Herman I. M., Taylor D. L. Correlated distribution of actin, myosin, and microtubules at the leading edge of migrating Swiss 3T3 fibroblasts. Cell Motil Cytoskeleton. 1989;14(4):527–543. doi: 10.1002/cm.970140410. [DOI] [PubMed] [Google Scholar]
  8. 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]
  9. Cortese J. D., Schwab B., 3rd, Frieden C., Elson E. L. Actin polymerization induces a shape change in actin-containing vesicles. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5773–5777. doi: 10.1073/pnas.86.15.5773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cunningham C. C. Actin polymerization and intracellular solvent flow in cell surface blebbing. J Cell Biol. 1995 Jun;129(6):1589–1599. doi: 10.1083/jcb.129.6.1589. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cunningham C. C., Stossel T. P., Kwiatkowski D. J. Enhanced motility in NIH 3T3 fibroblasts that overexpress gelsolin. Science. 1991 Mar 8;251(4998):1233–1236. doi: 10.1126/science.1848726. [DOI] [PubMed] [Google Scholar]
  12. Dancker P., Löw I., Hasselbach W., Wieland T. Interaction of actin with phalloidin: polymerization and stabilization of F-actin. Biochim Biophys Acta. 1975 Aug 19;400(2):407–414. doi: 10.1016/0005-2795(75)90196-8. [DOI] [PubMed] [Google Scholar]
  13. De La Cruz E. M., Pollard T. D. Transient kinetic analysis of rhodamine phalloidin binding to actin filaments. Biochemistry. 1994 Dec 6;33(48):14387–14392. doi: 10.1021/bi00252a003. [DOI] [PubMed] [Google Scholar]
  14. DeBiasio R. L., LaRocca G. M., Post P. L., Taylor D. L. Myosin II transport, organization, and phosphorylation: evidence for cortical flow/solation-contraction coupling during cytokinesis and cell locomotion. Mol Biol Cell. 1996 Aug;7(8):1259–1282. doi: 10.1091/mbc.7.8.1259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. DeBiasio R. L., Wang L. L., Fisher G. W., Taylor D. L. The dynamic distribution of fluorescent analogues of actin and myosin in protrusions at the leading edge of migrating Swiss 3T3 fibroblasts. J Cell Biol. 1988 Dec;107(6 Pt 2):2631–2645. doi: 10.1083/jcb.107.6.2631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Egelman E. H. The structure of F-actin. J Muscle Res Cell Motil. 1985 Apr;6(2):129–151. doi: 10.1007/BF00713056. [DOI] [PubMed] [Google Scholar]
  17. Erickson C. A. The deformability of BHK cells and polyoma virus-transformed BHK cells in relation to locomotory behaviour. J Cell Sci. 1980 Aug;44:187–200. doi: 10.1242/jcs.44.1.187. [DOI] [PubMed] [Google Scholar]
  18. 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. 1988;11(4):235–247. doi: 10.1002/cm.970110403. [DOI] [PubMed] [Google Scholar]
  19. Forscher P., Smith S. J. Actions of cytochalasins on the organization of actin filaments and microtubules in a neuronal growth cone. J Cell Biol. 1988 Oct;107(4):1505–1516. doi: 10.1083/jcb.107.4.1505. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Friederich E., Gouin E., Hellio R., Kocks C., Cossart P., Louvard D. Targeting of Listeria monocytogenes ActA protein to the plasma membrane as a tool to dissect both actin-based cell morphogenesis and ActA function. EMBO J. 1995 Jun 15;14(12):2731–2744. doi: 10.1002/j.1460-2075.1995.tb07274.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Giuliano K. A., Taylor D. L. Measurement and manipulation of cytoskeletal dynamics in living cells. Curr Opin Cell Biol. 1995 Feb;7(1):4–12. doi: 10.1016/0955-0674(95)80038-7. [DOI] [PubMed] [Google Scholar]
  22. Harris A., Dunn G. Centripetal transport of attached particles on both surfaces of moving fibroblasts. Exp Cell Res. 1972 Aug;73(2):519–523. doi: 10.1016/0014-4827(72)90084-5. [DOI] [PubMed] [Google Scholar]
  23. Hartwig J. H., Shevlin P. The architecture of actin filaments and the ultrastructural location of actin-binding protein in the periphery of lung macrophages. J Cell Biol. 1986 Sep;103(3):1007–1020. doi: 10.1083/jcb.103.3.1007. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Heacock C. S., Eidsvoog K. E., Bamburg J. R. The influence of contact-inhibited growth and of agents which alter cell morphology on the levels of G- and F-actin in cultured cells. Exp Cell Res. 1984 Aug;153(2):402–412. doi: 10.1016/0014-4827(84)90609-8. [DOI] [PubMed] [Google Scholar]
  25. Hill T. L. Microfilament or microtubule assembly or disassembly against a force. Proc Natl Acad Sci U S A. 1981 Sep;78(9):5613–5617. doi: 10.1073/pnas.78.9.5613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Huang Z. J., Haugland R. P., You W. M., Haugland R. P. Phallotoxin and actin binding assay by fluorescence enhancement. Anal Biochem. 1992 Jan;200(1):199–204. doi: 10.1016/0003-2697(92)90299-m. [DOI] [PubMed] [Google Scholar]
  27. Lanni F., Waggoner A. S., Taylor D. L. Structural organization of interphase 3T3 fibroblasts studied by total internal reflection fluorescence microscopy. J Cell Biol. 1985 Apr;100(4):1091–1102. doi: 10.1083/jcb.100.4.1091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lanni F., Ware B. R. Detection and characterization of actin monomers, oligomers, and filaments in solution by measurement of fluorescence photobleaching recovery. Biophys J. 1984 Jul;46(1):97–110. doi: 10.1016/S0006-3495(84)84002-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lin C. H., Espreafico E. M., Mooseker M. S., Forscher P. Myosin drives retrograde F-actin flow in neuronal growth cones. Neuron. 1996 Apr;16(4):769–782. doi: 10.1016/s0896-6273(00)80097-5. [DOI] [PubMed] [Google Scholar]
  30. Luby-Phelps K., Castle P. E., Taylor D. L., Lanni F. Hindered diffusion of inert tracer particles in the cytoplasm of mouse 3T3 cells. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4910–4913. doi: 10.1073/pnas.84.14.4910. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Mermall V., Post P. L., Mooseker M. S. Unconventional myosins in cell movement, membrane traffic, and signal transduction. Science. 1998 Jan 23;279(5350):527–533. doi: 10.1126/science.279.5350.527. [DOI] [PubMed] [Google Scholar]
  32. Mogilner A., Oster G. Cell motility driven by actin polymerization. Biophys J. 1996 Dec;71(6):3030–3045. doi: 10.1016/S0006-3495(96)79496-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Mooseker M. S., Pollard T. D., Wharton K. A. Nucleated polymerization of actin from the membrane-associated ends of microvillar filaments in the intestinal brush border. J Cell Biol. 1982 Oct;95(1):223–233. doi: 10.1083/jcb.95.1.223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Nishida E., Iida K., Yonezawa N., Koyasu S., Yahara I., Sakai H. Cofilin is a component of intranuclear and cytoplasmic actin rods induced in cultured cells. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5262–5266. doi: 10.1073/pnas.84.15.5262. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Peskin C. S., Odell G. M., Oster G. F. Cellular motions and thermal fluctuations: the Brownian ratchet. Biophys J. 1993 Jul;65(1):316–324. doi: 10.1016/S0006-3495(93)81035-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Petersen N. O., McConnaughey W. B., Elson E. L. Dependence of locally measured cellular deformability on position on the cell, temperature, and cytochalasin B. Proc Natl Acad Sci U S A. 1982 Sep;79(17):5327–5331. doi: 10.1073/pnas.79.17.5327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Pollard T. D. Rate constants for the reactions of ATP- and ADP-actin with the ends of actin filaments. J Cell Biol. 1986 Dec;103(6 Pt 2):2747–2754. doi: 10.1083/jcb.103.6.2747. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Post P. L., DeBiasio R. L., Taylor D. L. A fluorescent protein biosensor of myosin II regulatory light chain phosphorylation reports a gradient of phosphorylated myosin II in migrating cells. Mol Biol Cell. 1995 Dec;6(12):1755–1768. doi: 10.1091/mbc.6.12.1755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Rotsch C., Jacobson K., Radmacher M. Dimensional and mechanical dynamics of active and stable edges in motile fibroblasts investigated by using atomic force microscopy. Proc Natl Acad Sci U S A. 1999 Feb 2;96(3):921–926. doi: 10.1073/pnas.96.3.921. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Small J. V., Herzog M., Anderson K. Actin filament organization in the fish keratocyte lamellipodium. J Cell Biol. 1995 Jun;129(5):1275–1286. doi: 10.1083/jcb.129.5.1275. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Small J. V. Organization of actin in the leading edge of cultured cells: influence of osmium tetroxide and dehydration on the ultrastructure of actin meshworks. J Cell Biol. 1981 Dec;91(3 Pt 1):695–705. doi: 10.1083/jcb.91.3.695. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Smith S. J. Neuronal cytomechanics: the actin-based motility of growth cones. Science. 1988 Nov 4;242(4879):708–715. doi: 10.1126/science.3055292. [DOI] [PubMed] [Google Scholar]
  43. Spudich J. A., Watt S. The regulation of rabbit skeletal muscle contraction. I. Biochemical studies of the interaction of the tropomyosin-troponin complex with actin and the proteolytic fragments of myosin. J Biol Chem. 1971 Aug 10;246(15):4866–4871. [PubMed] [Google Scholar]
  44. Stossel T. P. The machinery of cell crawling. Sci Am. 1994 Sep;271(3):54-5, 58-63. doi: 10.1038/scientificamerican0994-54. [DOI] [PubMed] [Google Scholar]
  45. Theriot J. A., Mitchison T. J. Comparison of actin and cell surface dynamics in motile fibroblasts. J Cell Biol. 1992 Oct;119(2):367–377. doi: 10.1083/jcb.119.2.367. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Wang F. S., Wolenski J. S., Cheney R. E., Mooseker M. S., Jay D. G. Function of myosin-V in filopodial extension of neuronal growth cones. Science. 1996 Aug 2;273(5275):660–663. doi: 10.1126/science.273.5275.660. [DOI] [PubMed] [Google Scholar]
  47. Wang Y. L. Exchange of actin subunits at the leading edge of living fibroblasts: possible role of treadmilling. J Cell Biol. 1985 Aug;101(2):597–602. doi: 10.1083/jcb.101.2.597. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Wehland J., Osborn M., Weber K. Phalloidin-induced actin polymerization in the cytoplasm of cultured cells interferes with cell locomotion and growth. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5613–5617. doi: 10.1073/pnas.74.12.5613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Witke W., Sharpe A. H., Hartwig J. H., Azuma T., Stossel T. P., Kwiatkowski D. J. Hemostatic, inflammatory, and fibroblast responses are blunted in mice lacking gelsolin. Cell. 1995 Apr 7;81(1):41–51. doi: 10.1016/0092-8674(95)90369-0. [DOI] [PubMed] [Google Scholar]

Articles from Biophysical Journal are provided here courtesy of The Biophysical Society

RESOURCES