Ras oncogenes encode 21-kDa GTP-binding proteins that are capable of transforming immortalized cells in culture. Ras proteins are bound to the inner face of the plasma membrane by their C-terminal extremity and are thought to transmit their mitogenic signals via an "effector" domain spanning amino acids 32-42. Two ras-related human genes rap1A and rap1B encode 95% homologous 21-kDa proteins that share with Ras p21 the same effector domain and a similar C-terminal Cys-Ali-Ali-Xaa sequence (where Ali is an aliphatic amino acid; also known as a CAAX sequence). The product of the rap1A gene is identical to that of the Krev-1 cDNA, whose overexpression is capable of reverting the phenotype of Ki-ras-transformed NIH 3T3 cells. Antibodies that do not cross-react with Ras and other Ras-related proteins were obtained by immunizing rabbits with a peptide encompassing residues 121-137 of Rap1 proteins. These antibodies were used to investigate the subcellular localization of Rap1 proteins by indirect immunofluorescence and fractionation techniques. Rap1 proteins were found to be tightly bound to cellular membranes. They did not colocalize with Ras proteins on the plasma membrane and were discovered to be associated with the Golgi complex.