Actin from Thyone sperm assembles on only one end of an actin filament: a behavior regulated by profilin.

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Abstract
Thyone sperm was demembranated with Triton X-100 and, after washing, extracted with 30 mM Tris at ph 8.0 and 1 mM MgCl2. After the insoluble contaminents were removed by centrifugation, the sperm extract was warmed to 22.degree. C. Actin filaments rapidly assembled and aggregated into bundles when KCl was added to the extract. When preformed actin filaments, i.e., the acrosomal filament bundles of Limulus sperm, were added to the extract, the actin monomers rapidly assembled on these filaments. What was unexpected was that assembly took place on only 1 end of the bundle, the end corresponding to the preferred end for monomer addition. The absence of growth on the nonpreferred end was not due to the presence of a capper because exogenously added actin readily assembled on both ends. The sperm extract by sodium dodecyl sulfate gel electrophoresis was also analyzed. Two major proteins were present in a 1:1 molar ratio: actin and a 12,500-dalton protein whose apparent isoelectric point was 8.4. The 12,500-dalton protein was purified by DEAE chromatography. It is profilin because of its size, isoelectric point, molar ratio to actin, inability to bind to DEAE and its effect on actin assembly. When profilin was added to actin in the presence of Limulus bundles, addition of monomers on the nonpreferred end of the bundle was inhibited, even though actin by itself assembled on both ends. Using the Limulus bundles as nuclei, the critical concentration for assembly off each end of the filament was determined and the Kd was estimated for the profilin-actin complex (.apprx. 10 .mu.M). A model is presented to explain how profilin may regulate the extension of the Thyone acrosomal process in vivo. The profilin-actin complex can add to only the preferred end of the filament bundle. Once the actin monomer is bound to the filament, the profilin is released and is available to bind to additional actin monomers. This mechanism accounts for the rapid rate of filament elongation in the acrosomal process in vivo.