Expression of the c‐erb B‐2 protein in normal and transformed cells

Abstract

Two synthetic peptides from the predicted sequence of the human c‐erb B‐2 protein were synthesized and used to raise antisera in rabbits. The sequences chosen were identical to those in the homologous rat c‐neu protein. The antibodies produced reacted with the immunizing peptides in ELISA but showed little or no cross‐reaction with the partly homologous peptides found in equivalent positions in the human EGF receptor. Both antipeptide antibodies, and a monoclonal antibody (MAb) specific for the rat new protein, immuno‐precipitated a 185‐kDa protein from 35 S‐methionine‐labelled lysates prepared from a rat cell line known to express high levels of the c‐neu protein. The antipeptide antibodies also recognized a protein of the same size in Western blots. In addition, both antipeptide antibodies immunoprecipitated a 190‐kDa protein from labelled cell lysates prepared from human and monkey cells. Antibodies to one of the peptides, which showed no detectable cross‐reaction with human, rat or monkey EGF receptor, were used to examine the expression of the c‐erb B‐2 protein on a variety of cultured cell types. Eleven transformed, I non‐established and 2 immortalized cell types were examined by immunoprecipitation for their level of expression of the c‐erb B‐2 protein and of the EGF receptor. The numbers of EGF receptors varied widely between different cell lines, whereas the level of the c‐erb B‐2 protein, which was found on all of the cell types examined, was more constant. The number of c‐erb B‐2 molecules present was estimated by autoradiography to be about 100,000 per cell. The antibodies were then used to examine the location and level of expression of the human c‐erb B‐2 and rat c‐neu proteins in normal tissues. Immunohistochemical staining showed that the c‐erb B‐2 protein was highly expressed in rat kidney proximal tubules and loop of Henle. The c‐enb B‐2 protein was also present on normal human epithelial cells but in some cases with a different distribution to that of the EGF receptor.