DYNAMICS OF FILAMENTOUS ACTIN ORGANIZATION IN THE SEA-URCHIN EGG CORTEX DURING EARLY CLEAVAGE DIVISIONS - IMPLICATIONS FOR THE MECHANISM OFCYTOKINESIS

We have used confocal laser scanning microscopy in conjunction with BO DIPY-phallacidin staining of filamentous actin to investigate changes in the quantity and organization of cortical actin during the first tw o cell cycles following fertilization in eggs of the sea urchin Strong lycoentrotus purpuratus. Quantification of fluorescent phallacidin sta ining reveals that the amount of filamentous actin (F-actin) in the co rtex undergoes cyclical increases and decreases during early cleavage divisions, peaking near the beginning of the cell cycle and decreasing to a minimum at cytokinesis. Changes in the content of cortical F-act in are accompanied by the growth and disappearance of rootlet-like bun dles of actin filaments which extend from the bases of microvilli that cover the surface of the egg. Actin rootlets reach their maximum degr ee of development by 20 min postfertilization, and then gradually decr ease in number and length over the next 40 min. Small actin rootlets p ersist until cleavage, disappear during cytokinesis, and reform follow ing division. The formation of actin rootlets requires cytoplasmic alk alization and is inhibited by cytochalasin D. Cytochalasin D washout e xperiments demonstrate that assembly of the cortical actin cytoskeleto n can be blocked until 5 min before the onset of cleavage and still al low normal cytokinesis. These results illustrate the dynamic nature of cortical. actin organization during early development and demonstrate that cytokinesis occurs at the point of minimum cortical F-actin cont ent. They further demonstrate that cytokinesis can occur in embryos in which the normal developmental sequence of changes in cortical actin organization has been blocked by treatment with cytochalasin D, sugges ting that these changes do not function in the establishment of the co ntractile apparatus for cytokinesis, but rather serve other developmen tal functions. (C) 1997 Wiley-Liss, Inc.