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These movies accompany the manuscript Clarke et al., Mol. Biol. Cell 17, 4866-4875 (2006), which is freely accessible at the journal website, http://www.molbiolcell.org. The movies illustrate actin-mediated phagosome movement induced by compressing the cell, bringing a previously-ingested phagosome into contact with the cortical actin layer.

Movie 1A.

Clarke_Movie-01A.mov, 256x256 pixels, 0.2MB compressed

 

 Pressure-induced actin accumulation and phagosome movement viewed by TIRF microscopy. The phagosomes (containing TRITC-labeled yeast) moved in changing directions according to the site of strongest actin deposition. (Figure 1A)
 

Movie 1B.

Clarke_Movie-01B.mov, 113x108 pixels, 0.1MB compressed

 Although phagosome movement was commonly preceded by an annular accumulation of actin, sometimes a single focus of actin was observed.
 

Movie 2.

Clarke_Movie-02.mov, 320x301 pixels, 5.0MB compressed

 A TIRF view of the induction of phagosome movement in several adjoining cells by gradual drying of an agar overlay. The phagosomes exhibited multiple cycles of rocketing, with MyoB-GFP accumulating during each pause. (Figure 1B)
 

Movie 3.

Clarke_Movie-03.mov, 345x355 pixels, 2.5MB compressed

 For moving phagosomes, GFP-Arp3 and LimEΔ exhibited almost complete overlap, although the mRFP-LimEΔ signal was slightly enriched close to the phagosome in the deeper focal plane shown here. (Figure 3C)
 

Movie 4.

Clarke_Movie-04.mov, 379x341 pixels, 3.7MB compressed

 Each time a phagosome paused, a ring of GFP-Arp3 appeared at the plasma membrane. When the phagosome moved away, the ring was left behind and eventually disappeared. (Figure 4A)
 

Movie 5A.

Clarke_Movie-05A.mov, 543x243 pixels, 2.6MB compressed

 Back-and-forth phagosome movement in cells expressing GFP-Arp3.
 

Movie 5B.

Clarke_Movie-05B.mov, 318x325 pixels, 4.9MB compressed

 Continuing view of a cell from Movie 5A, showing conversion of back-and-forth movement to pinwheeling. The focus was shifted from the plasma membrane to the cell center and back, revealing GFP-Arp3 only near the plasma membrane. (Figure 4B)
 

Movie 6.

Clarke_Movie-06.mov, 391x456 pixels, 5.2MB compressed

 Rocketing phagosome in a cell expressing mRFP-LimEΔ and coronin-GFP. Each change of direction was accompanied by new actin filaments, which were labeled first by mRFP-LimEΔ, then by coronin-GFP, and finally disappeared. (Figure 5)
 

Movie 7.

Clarke_Movie-07.mov, 404x243 pixels, 4.8MB compressed

 VatM-GFP was present in the membrane of these rocketing phagosomes, indicating that the phagosomes were in the acidic stage of endocytic transit. (Figure 6A)
 

Movie 8.

Clarke_Movie-08a.mov, 297x339 pixels, 4.4MB compressed

 This rocketing phagosome made three circuits through the cytoplasm during the 232-second time series, traveling at an average velocity of 23 μm/min. It rammed and distorted the nucleus (red) on each circuit. (Figure 7A)
 

Movie 9.

Clarke_Movie-09.mov, 306x373 pixels, 5.2MB compressed

 This rocketing phagosome, constrained and guided by microtubules, often collided with the centrosome and nucleus, exhibiting sufficient force to displace the nucleus and straighten the microtubules. (Figure 7B)
 

Supplemental Movie 1.

Clarke_Suppl-Movie.mov, 247x247 pixels, 4.9MB compressed

 Phagosome behavior in a myoB- mutant. When the myoB- cell was compressed by an agarose overlay, actin accumulated about the two yeast-containing phagosomes, and one of them began to move. However, the moving phagosome exhibited limited displacement and exerted little apparent force. This movie, like Movie 9, is shown at 24x actual speed.
last updated: December 12, 2006
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