THE ROLE OF SPHINGOLIPIDS IN THE MAINTENANCE OF FIBROBLAST MORPHOLOGY- THE INHIBITION OF PROTRUSIONAL ACTIVITY, CELL SPREADING, AND CYTOKINESIS INDUCED BY FUMONISIN B-1 CAN BE REVERSED BY GANGLIOSIDE GM(3)

Previous studies demonstrated that inhibition of sphingolipid synthesi s by the mycotoxin fumonisin B-1 (FB1) disrupts axonal growth in cultu red hippocampal neurons (Harel, R., and Futerman, A. H. (1993) J. Biol . Chem. 268, 14476-14481) by affecting the formation or stabilization of axonal branches (Schwarz, A., Rapaport, E., Hirschberg, K., and Fut erman, A.H. (1995) J. Biol. Chem. 270, 10990-10998). We now demonstrat e that long term incubation with FB1 affects fibroblast morphology and proliferation. Incubation of Swiss 3T3 cells with FB1 resulted in a d ecrease in synthesis of ganglioside GM(3), the major glycosphingolipid in 3T3 fibroblasts and of sphingomyelin. The projected cell area of F B1-treated cells was similar to 45% less than control cells. FB1 had n o affect on the organization of microtubules or intermediate filaments , but fewer actin-rich stress fibers were observed, and there was a lo ss of actin-rich lamellipodia at the leading edge. Three other process es involving the actin cytoskeleton, cytokinesis, microvilli formation , and the formation of long processes induced by protein kinase inhibi tors, were all disrupted by FB1. All the effects of FB1 on cell morpho logy could be reversed by addition of ganglioside GM(3) even in the pr esence of FB1, whereas the bioactive intermediates, sphinganine, sphin gosine, and ceramide, were without effect. Finally, FB1 blocked cell p roliferation and DNA synthesis in a reversible manner, although gangli oside GM(3) could not reverse the effects of FB1 on cell proliferation . Together, these data suggest that ongoing sphingolipid synthesis is required for the assembly of both new membrane and of the underlying c ytoskeleton.