Immunulocalization of the gamma isoform of nonmuscle actin in cultured cells

Abstract

In many vertebrate nonmuscle cells, the microfilament subunit protein, actin, exists as two isoforms, called β and γ, whose sequences differ only in their amino-terminal regions. We have prepared a peptide antibody specifically reactive with the aminoterminal sequence of γ actin. This antibody reacted with nonmuscle actin as determined by Western blots of SDS gels, and reacted with the γ, but not the β, nonmuscle actin isoform as shown by Western blots of isoelectric focusing gels. In immunofluorescence experiments, the γ peptide antibody stained microfilaments bundles, ruffled edges, and the contractile ring of a variety of cultured cells, including mouse L cells, which have previously been reported to contain only the β actin isoform (Sakiyama, S., S. Fujimura, and H. Sakiyama, 1981, J. Biol. Chem., 256:31-33). Double immunofluorescence experiments using the γ peptide antibody and an antibody reactive with all actin isoforms revealed no differences in isoform localization. Thus, at the level of resolution of light microscopy, we have detected the γ actin isoform in all microfilament-containing structures in cultured cells, and have observed no subcellular sorting of the non-muscle actin isoforms.