Cingulin interacts with F-actin in vitro

Cingulin, a M-r 140-160 kDa protein of the cytoplasmic plaque of epithelial tight junctions (TJ), interacts in vitro with TJ proteins and myosin. Here we investigated cingulin interaction with actin, using His-tagged, full-le ngth Xenopus laevis cingulin expressed in insect cells, and glutathione S-t ransferase (GST) fusion proteins of fragments of cingulin expressed in bact eria. Purified full-length cingulin co-pelleted with F-actin after high spe ed centrifugation, and promoted the sedimentation of F-actin under low spee d centrifugation, suggesting that cingulin is an actin-cross-linking protei n. The actin interaction of GST fusion proteins containing fragments of Xen opus cingulin suggested that the F-actin binding site is between residues 1 01 and 294. (C) 2001 Federation of European Biochemical Societies. Publishe d by Elsevier Science B.V. All rights reserved.