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The 21-kDa GTP-binding Yptl protein (Yptlp) is required for protein transport from the endoplasmic reticulum to the Golgi complex in yeast extracts. Yptl antibodies block transport; this inhibition is alleviated by competition with excess purified Yptlp produced in bacteria. Fur-

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Abstract. F-actin affinity chromatography and immunological techniques are used to identify actin-binding proteins in purified Dictyostelium discoideum plasma membranes. A 17-kD integral glycoprotein (gpl7) consistently elutes from F-actin columns as the major actin-binding protein under a variety of experimental conditions. The actin-binding activity of gpl7 is identical to that of intact plasma membranes: it resists extraction with 0.1 N NaOH, 1 mM dithiothreitol (DTT); it is sensitive to ionic conditions; it is stable over a wide range of pH; and it is eliminated by proteolysis, denaturation with heat, or treatment with DTT and N-ethylmaleimide. gpl7 may be responsible for much of the actin-binding activity of plasma membranes since monovalent antibody fragments (Fab) directed primarily against gpl7 inhibit actin-membrane binding by 96% in sedimentation assays. In contrast, Fab directed against cell surface determinants inhibit binding by only 0-10%. The actin-binding site of gpl7 appears to be located on the cytoplasmic surface of the membrane since Fab against this protein continue to inhibit 96 % of actin-membrane binding even after extensive adsorption against cell surfaces. gpl7 is abundant in the plasma membrane, constituting 0.4-1.0% of the total membrane protein. A transmembrane orientation of gpl7 is suggested since, in addition to the cytoplasmic localization of the actinbinding site, extracellular determinants of gpl7 are identified, gpl7 is surface-labeled by sulfo-N-hydroxysuccinimido-biotin, a reagent that cannot penetrate the cell membrane. Also, gpl7 is glycosylated since it is specifically bound by the lectin, concanavalin A. We propose that gpl7 is a major actin-binding protein that is important for connecting the plasma membrane to the underlying microfilament network. Therefore, we have named this protein "ponticulin" from the Latin word, ponticulus, which means small bridge.