Reorganization of actin cytoskeleton at the growing end of the cleavage furrow of Xenopus egg during cytokinesis.

We studied reorganization of actin-myosin cytoskeleton at the growing ends of the cleavage furrow of Xenopus eggs in order to understand how the contractile ring is formed during cytokinesis. Reorganization of F-actin structures during the furrow formation was demonstrated by rhodamine-phalloidin staining of the cleavage furrow and by time-lapse scanning with laser scanning microscopy of F-actin structures in the cleavage furrow of live eggs to which rhodamine-G-actin had been injected. Actin filaments assemble to form small clusters that we call 'F-actin patches' at the growing end of the furrow. In live recordings, we observed emergence and rapid growth of F-actin patches in the furrow region. These patches then align in tandem, elongate and fuse with each other to form short F-actin bundles. The short bundles then form long F-actin bundles that compose the contractile ring. During the furrow formation, a cortical movement towards the division plane occurs at the growing ends of the furrow, as shown by monitoring wheatgerm agglutinin-conjugated fluorescent beads attached to the egg surface. As a result, wheatgerm agglutinin-binding sites accumulate and form 'bleb-like' structures on the surface of the furrow region. The F-actin patch forms and grows underneath this structure. The slope of F-actin accumulation in the interior region of the furrow exceeds that of accumulation of the cortex transported by the cortical movement. In addition, rhodamine-G-actin microinjected at the growing end is immediately incorporated into the F-actin patches. These data, together with the rapid growth of F-actin patches in the live image, suggest that actin polymerization occurs in the contractile ring formation. Distribution of myosin II in the cleavage furrow was also examined by immunofluorescence microscopy. Myosin II assembles as spots at the growing end underneath the bleb-like structure. It was suggested that myosin is transported and accumulates as spots by way of the cortical movement. F-actin accumulates at the position of the myosin spot a little later as the F-actin patches. The myosin spots and the F-actin patches are then simultaneously reorganized to form the contractile ring bundles