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. 2000 Jul;79(1):191–201. doi: 10.1016/S0006-3495(00)76283-7

Probing f-actin flow by tracking shape fluctuations of radial bundles in lamellipodia of motile cells.

G Danuser 1, R Oldenbourg 1
PMCID: PMC1300925  PMID: 10866947

Abstract

We examined the dynamics of radial actin bundles based on time-lapse movies of polarized light images of living neuronal growth cones. Using a highly sensitive computer vision algorithm for tracking, we analyzed the small shape fluctuations of radial actin bundles that otherwise remained stationary in their positions in the growth cone lamellipodium. Using the tracking software, we selected target points on radial bundles and measured both the local bundle orientations and the lateral displacements between consecutive movie frames. We found that the local orientation and the lateral displacement of a target point are correlated. The correlation can be explained using a simple geometric relationship between the lateral travel of tilted actin bundles and the retrograde flow of f-actin structures. Once this relationship has been established, we have turned the table and used the radial bundles as probes to measure the velocity field of f-actin flow. We have generated a detailed map of the complex retrograde flow pattern throughout the lamellipodium. Such two-dimensional flow maps will give new insights into the mechanisms responsible for f-actin-mediated cell motility and growth.

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Selected References

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  1. Danuser G., Tran P. T., Salmon E. D. Tracking differential interference contrast diffraction line images with nanometre sensitivity. J Microsc. 2000 Apr;198(Pt 1):34–53. doi: 10.1046/j.1365-2818.2000.00678.x. [DOI] [PubMed] [Google Scholar]
  2. Gelles J., Schnapp B. J., Sheetz M. P. Tracking kinesin-driven movements with nanometre-scale precision. Nature. 1988 Feb 4;331(6155):450–453. doi: 10.1038/331450a0. [DOI] [PubMed] [Google Scholar]
  3. Gittes F., Mickey B., Nettleton J., Howard J. Flexural rigidity of microtubules and actin filaments measured from thermal fluctuations in shape. J Cell Biol. 1993 Feb;120(4):923–934. doi: 10.1083/jcb.120.4.923. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Ishijima A., Doi T., Sakurada K., Yanagida T. Sub-piconewton force fluctuations of actomyosin in vitro. Nature. 1991 Jul 25;352(6333):301–306. doi: 10.1038/352301a0. [DOI] [PubMed] [Google Scholar]
  5. Katoh K., Hammar K., Smith P. J., Oldenbourg R. Arrangement of radial actin bundles in the growth cone of Aplysia bag cell neurons shows the immediate past history of filopodial behavior. Proc Natl Acad Sci U S A. 1999 Jul 6;96(14):7928–7931. doi: 10.1073/pnas.96.14.7928. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Katoh K., Hammar K., Smith P. J., Oldenbourg R. Birefringence imaging directly reveals architectural dynamics of filamentous actin in living growth cones. Mol Biol Cell. 1999 Jan;10(1):197–210. doi: 10.1091/mbc.10.1.197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Lin C. H., Forscher P. Growth cone advance is inversely proportional to retrograde F-actin flow. Neuron. 1995 Apr;14(4):763–771. doi: 10.1016/0896-6273(95)90220-1. [DOI] [PubMed] [Google Scholar]
  8. 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]
  9. Oldenbourg R. A new view on polarization microscopy. Nature. 1996 Jun 27;381(6585):811–812. doi: 10.1038/381811a0. [DOI] [PubMed] [Google Scholar]
  10. Oldenbourg R., Katoh K., Danuser G. Mechanism of lateral movement of filopodia and radial actin bundles across neuronal growth cones. Biophys J. 2000 Mar;78(3):1176–1182. doi: 10.1016/S0006-3495(00)76675-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Oldenbourg R., Mei G. New polarized light microscope with precision universal compensator. J Microsc. 1995 Nov;180(Pt 2):140–147. doi: 10.1111/j.1365-2818.1995.tb03669.x. [DOI] [PubMed] [Google Scholar]
  12. Oldenbourg R., Salmon E. D., Tran P. T. Birefringence of single and bundled microtubules. Biophys J. 1998 Jan;74(1):645–654. doi: 10.1016/S0006-3495(98)77824-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Small J., Rottner K., Hahne P., Anderson K. I. Visualising the actin cytoskeleton. Microsc Res Tech. 1999 Oct 1;47(1):3–17. doi: 10.1002/(SICI)1097-0029(19991001)47:1<3::AID-JEMT2>3.0.CO;2-2. [DOI] [PubMed] [Google Scholar]
  14. Svoboda K., Schmidt C. F., Schnapp B. J., Block S. M. Direct observation of kinesin stepping by optical trapping interferometry. Nature. 1993 Oct 21;365(6448):721–727. doi: 10.1038/365721a0. [DOI] [PubMed] [Google Scholar]
  15. Theriot J. A., Mitchison T. J. Actin microfilament dynamics in locomoting cells. Nature. 1991 Jul 11;352(6331):126–131. doi: 10.1038/352126a0. [DOI] [PubMed] [Google Scholar]
  16. Wang Y. L. Reorganization of actin filament bundles in living fibroblasts. J Cell Biol. 1984 Oct;99(4 Pt 1):1478–1485. doi: 10.1083/jcb.99.4.1478. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. 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. 1998 Nov 5;8(22):1227–1230. doi: 10.1016/s0960-9822(07)00515-5. [DOI] [PubMed] [Google Scholar]
  18. Yin H., Landick R., Gelles J. Tethered particle motion method for studying transcript elongation by a single RNA polymerase molecule. Biophys J. 1994 Dec;67(6):2468–2478. doi: 10.1016/S0006-3495(94)80735-0. [DOI] [PMC free article] [PubMed] [Google Scholar]

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