Immunofluorescence and immunoblotting techniques were used to study the presence and distribution of
vimentin and
keratin type intermediate filaments, actin, and
vinculin (130 kD
protein) during
retinoic acid (RA)-induced differentiation of F9
embryonal carcinoma (EC) cells. The undifferentiated F9 cells regularly expressed
vimentin, usually concentrated close to the nucleus, but not
keratin. Actin appeared as short intracellular filaments and as spikes at the edges of the colonies, together with some diffuse cytoplasmic staining. F9 cells also showed a weak, diffuse cytoplasmic
vinculin-specific fluorescence in addition to occasional small focal
vinculin patches at the edges of the cell colonies. RA treatment led into a series of changes in the cytoskeletal organization of F9 cells. These changes were initiated by the appearance of distinct
vinculin plaques and followed by formation of actin stress fibers and by profound changes in the organization of
vimentin in the flattening cells. RA treatment finally led to the appearance and co-expression of
keratin fibrils in many of the
vimentin-containing F9 cells. This sequence of changes suggests that the
vinculin-containing adhesion plaques may be important in the mechanism of RA-induced differentiation of EC cells.