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Daniel Zicha,1Ian M. Dobbie,2Mark R. Holt,3James Monypenny,1Daniel Y. H. Soong,3Colin Gray,1Graham A. Dunn3*
Transformed rat fibroblasts expressing two variants of green
fluorescent protein, each fused to -actin, were used to studyactin
dynamics during cell protrusion. The recently developedFLAP
(fluorescence localization after photobleaching) method permitsthe
tracking of one fluorophore after localized photobleachingby using the
other as a colocalized reference. Here, by visualizingthe ratio of
bleached to total molecules, we found that actinwas delivered to
protruding zones of the leading edge of the cellat speeds that
exceeded 5 micrometers per second. Monte Carlomodeling confirmed that
this flow cannot be explained by diffusionand may involve active
transport.
1 Light Microscopy, Cancer Research UK,
Lincoln's Inn Fields Laboratories, London WC2A 3PX, UK.
2 Genome Stability Laboratory, Department of
Biochemistry, National University of Ireland, Galway, Ireland.
3 The Randall Centre, New Hunt's House, Guy's
Campus, King's College London, London SE1 1UL, UK.
*
To whom correspondence should be addressed. E-mail:
graham.a.dunn{at}kcl.ac.uk
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-actin, were used to study actin
dynamics during cell protrusion. The recently developed FLAP
(fluorescence localization after photobleaching) method permits the
tracking of one fluorophore after localized photobleaching by using the
other as a colocalized reference. Here, by visualizing the ratio of
bleached to total molecules, we found that actin was delivered to
protruding zones of the leading edge of the cell at speeds that
exceeded 5 micrometers per second. Monte Carlo modeling confirmed that
this flow cannot be explained by diffusion and may involve active
transport.
