'Contact regions' are plasma membrane domains derived from areas of intercellular contact between aggregating Dictyostelium amebae (H.M. Ingalls et al. (1986). Proc. Nat. Acad. Sci. USA 83, 4779). Purified contact regions contain a prominent actin-binding protein with an M(r) of 34,000. Immunoblotting with monoclonal antibodies identifies this polypeptide as a 34,000 M(r) actin-bundling protein (known as 30 kDa protein), previously shown to be enriched in filopodia (M. Fechheimer (1987). J. Cell Biol. 104, 1539). About four times more 30 kDa protein by mass is associated with contact regions than is found in total plasma membranes isolated from aggregating cells. In agreement with these observations, immunostaining of the 30 kDa protein in aggregating cells reveals a prominent localization along the plasma membrane at sites of intercellular contact. By contrast, alpha-actinin does not appear to be significantly enriched at sites of cell to cell contact. Binding experiments using purified plasma membranes, actin and 30 kDa protein indicate that the 30 kDa protein is associated with the plasma membrane primarily through interactions with actin filaments. Calcium ions are known to decrease the interaction of actin with 30 kDa protein in solution. Surprisingly, membrane-associated complexes of actin and the 30 kDa protein are much less sensitive to dissociation by micromolar levels of free calcium ions than are complexes in solutions lacking membranes.(ABSTRACT TRUNCATED AT 250 WORDS)