Formation of filopodia in coelomocytes: localization of fascin, a 58,000 dalton actin cross-linking protein.

Echinoderm coelomocytes or phagocytes purified in the petaloid stage will attach to a glass substrate and form large circumferential lamellIpodia. Hypotonic shock will induce quantitative transformation to a filopodial form. Differentiation of the peripheral cytoplasm begins at the cell edge, when phase dense rods composed of actin filaments start to form. These structures, which eventually form the cores of filopodia, continue to grow, lengthen and extend deeper into the cytoplasm. In the final stage, the plasma membrane retracts down around a core to form a filopodium. Antibody against a 58,000 dalton protein isolated from sea urchin egg actin gels has been used to study a rather striking redistribution of this protein in the peripheral cytoplasm of the coelomocyte during the transformation sequence. This protein is known to organize actin filaments in vitro into linear paracrystalline arrays with a distinct 11 nm banding pattern by forming cross-links between adjacent actin filaments. In the early stage of the transformation, indirect immunofluorescence indicates a random distribution of this protein in the circumferential lamellipodia. Organization is first seen at the cell edge, where fluorescent rods coincident with the phase-dense structures start to form. These rods lengthen, extend deeper into the cytoplasm and become more intensely fluorescent. After membrane retraction, cells with individual, intensely stained filopodia are visible. The known chemistry of the actin cross-linking protein (M_r = 58,000) and its redistribution during the transformation sequence are consistent with the idea that this protein functions to organize F actin into filopodial cores by cross-linking adjacent actin filaments. We have named this protein “fascin,” because it organizes actin filaments, both in vivo and in vitro, into linear arrays or fascicles. Antibody staining shows a second population of these actin cross-linking molecules localized in the perinuclear cytoplasm. In this region, fascin appears to function to maintain a stable circumnuclear cage structure which is part of the coelomocyte cytoskeleton.