Transformation of chicken embryo fibroblasts by avian retroviruses induces the tyrosine phosphorylation of a 34–39 kD cellular protein (p34). In vitro, p34 isolated from intestine interacts with F-actin in a Ca²⁺-dependent manner. We report here that, in the absence of Ca²⁺ chelators, three proteins co-purified with p34 extracted from a cytosolic or membrane fraction of chicken embryo fibroblasts; these two fractions account respectively for 10–20% and 50% of the total cellular p34. Isolated from the cytosoluble fraction of fibroblasts by sucrose gradient centrifugation and hydrophobic chromatography, p34 and the other proteins behaved as a homogeneous species upon non-denaturing gel electrophoresis, gel filtration, and CsCl density gradient centrifugation, thus indicating a strong association. Moreover, an analysis by electron microscopy following uranyl acetate staining revealed particles with a raspberry-like shape. This association was always disrupted by the calcium-chelating agent, EGTA.