Tolytoxin, a cytostatic, antifungal macrolide produced by blue-green algae of the genus Scytonema, is a potent, reversible inhibitor of cytokinesis in cultured mammalian cells. Treatment of KB cells with 2-16 nM tolytoxin results in profound morphological changes, beginning with the formation of zeiotic processes and culminating in nuclear protrusion. In L1210 cells, cytokinesis is inhibited by as little as 2 nM tolytoxin, while karyokinesis proceeds normally, resulting in polynucleation. Tolytoxin specifically disrupts microfilament organization in A10 cells, while having no apparent effect on microtubules or intermediate filaments. Tolytoxin inhibited actin polymerization in vitro and also caused the depolymerization or fragmentation of F-actin in vitro. Tolytoxin exhibits effects that closely resemble those of cytochalasin B but is effective at concentrations 1/50-1/1,000 that of cytochalasin B.