CA2-DEPENDENT INTERACTION OF S100A2 WITH MUSCLE AND NONMUSCLE TROPOMYOSINS()

Zero-length chemical crosslinking with 1-ethyl-3-[3-(dimethyl amino)pr opyl]carbodiimide (EDC) indicated an association of the Ca2+-binding p rotein S100A2 with tropomyosin (TM) in vitro. The mobility of the cros slinked product on SDS-PAGE gels indicated the formation of a 1:1 comp lex between S100A2 and TM and the interaction was Ca2+ dependent. Mono clonal antibodies were raised against S100A2 and used to determine its cellular localization in the porcine epithelial cell line LLC PK1. It was found that the localization of S100A2 depended on the differentia tion state of the cells, being absent from actin stress fibers in spar sely seeded cultures, but present in the actin-containing microvilli c haracteristic of differentiated cells. Immuneprecipitations of [S-35] methionine-labeled extracts using S100A2 as well as TM-specific antibo dies failed to co-precipitate TM and S100A2, indicating a transient as sociation between these two molecules in solution. Affinity chromatogr aphy of cell extracts on immobilized recombinant TMs, however, confirm ed the Ca2+-dependent interaction between S100A2 and both muscle TMs a s well as with high and low molecular mass nonmuscle TMs, suggesting t hat the binding site resides in one of the conserved regions of TM. Ou r data demonstrate the possible interaction of S100A2 with TM that is not bound to the microfilaments and indicate a differentiation-related function for S100A2 in LLC PK1 cells. The possible functional implica tions of this interaction are discussed.