p185, a product of the neu proto-oncogene, is a receptorlike protein associated with tyrosine kinase activity.

Abstract

The neu oncogene was originally identified in cell lines derived from rat neuroectodermal tumors. neu is related to but distinct from the c-erbB gene, which encodes the epidermal growth factor (EGF) receptor. neu encodes a protein, designated p185, that is serologically related to the EGF receptor. Identification of the normal homolog of p185 encoded by the neu proto-oncogene enabled us to compare the product of the neu proto-oncogene with the mutated version specified by the neu oncogene and with the EGF receptor. The normal form of p185 was structurally similar to its transforming counterpart, indicating that activation of the neu oncogene did not cause major structural alterations in the gene product. Both normal and transforming forms of p185 were associated with tyrosine kinase activity, supporting the idea that normal p185 functions as a growth factor receptor. p185 differed both structurally and functionally from the EGF receptor. p185 and the EGF receptor had distinct electrophoretic mobilities when synthesized under normal culture conditions or in the presence of tunicamycin. EGF did not stimulate increased turnover of p185 and did not bind quantitatively to p185. A number of other growth factors failed to stimulate degradation of p185 or tyrosine phosphorylation of p185 and are therefore unlikely to be ligands for p185.More than 20 different oncogenes have been implicated in virus-mediated tumorigenesis, and a number of these are likely to play a direct role in human cancer (reviewed in references 4 and 35). It is now evident that some of these oncogenes operate by disrupting pathways regulated by polypeptide growth factors (reviewed in references 23, 25, and 59). Thus the v-sis oncogene has been found to encode a protein which is structurally and functionally homologous to platelet-derived growth factor (PDGF) (13,15,30,48,64). v-sis apparently transforms cells by causing them to constitutively secrete a mitogenic factor to which these same cells respond. Two viral oncogenes, v-erbB and v-fms, are derived from cellular genes encoding growth factor receptors. v-erbB specifies a truncated version of the cellular epidermal growth factor receptor (EGFr) (16, 63), while v-fms specifies a protein related to the macrophage colony-stimulating factor 1 receptor (56). The two defective receptor proteins may transform cells by constitutively transmitting growth excitatory signals that are normally produced only in the presence of their respective ligands.We have been studying another oncogene, designated neu, which apparently specifies a mutated growth factor receptor. In contrast to v-erbB and v-fms, which were discovered as transforming genes of retroviruses, the neu oncogene was identified by transfection of DNA from chemically induced tumors (57). DNA from four ethyl-nitrosourea-induced rat neuroectodermal tumor cell lines induced foci when introduced by transfection into NIH 3T3 cell monolayers (57). Subsequent studies revealed that these four neuroectodermal tumor cell lines all contained activated version...

Expression of dominant-negative ErbB2 in the mammary gland of transgenic mice reveals a role in lobuloalveolar development and lactation

“…The ErbB2DIC protein was expressed in FR3T3 rat embryonic ®broblast cells transfected with MMTVErbB2DIC ( Figure 1b, lane 2) but not cells transfected with an empty vector ( Figure 1b, lane 1). ErbB2 is phosphorylated in response to EGF when coexpressed with EGFR (Stern et al, 1986;Stern and Kamps, 1988). To determine the e ects of ErbB2DIC expression on the transphosphorylation of ErbB2 in response to EGF, FR3T3 cell lines expressing ErbB2DIC or the MMTV vector alone were mock treated or treated with EGF.…”

“…Stable cell clones in 60 mm dishes were metabolically labeled, in the presence of 1 mM dexamethasone, by incubating for 16 h with 100 mCi of 35 Smethionine/cysteine in vitro cell labeling mix (Amersham). Labeled cells were lysed on ice in RIPA bu er (10 mM Tris, pH 7.4, 150 mM NaCl, 1% deoxycholate, 1% NP-40, 0.1% sodium dodecyl sulfate, with 1 mM phenylmethylsulfonyl uoride, and 1% aprotinin) and immunoprecipitations were performed by adding 1 mg of anti-c-neu Ab-4 (Oncogene Science) and 50 ml of preswollen protein A sepharose (Pharmacia) (Stern et al, 1986). Precipitated proteins were resolved by SDS ± PAGE on a 12% acrylamide gel and the resolving gel was processed in Opti¯uor (National Diagnostics) according to the manufacturer's instructions.…”

“…ErbB2 was immunoprecipitated from lysate containing 300 mg of protein and analysed by SDS-PAGE as described above. Phosphotyrosine Western blot was performed using mouse monoclonal antibody 4G10 (Upstate Biotechnology) diluted to 1 mg/ml using procedures described previously (Stern et al, 1986). The blot was stripped of primary antibody as described by the manufacturer (Pierce) and was reprobed for ErbB2 with Neu antibody C-18 (Santa Cruz) diluted to 0.2 mg/ml.…”

“…ErbB2 signaling can be activated, however, through heterodimer formation with other ligand-bound ErbB-family members. For example, the EGFR ligand, EGF does not bind to ErbB2 but does activate ErbB2 when ErbB2 is coexpressed with EGFR (Stern et al, 1986;Stern and Kamps, 1988). This is mediated by induction of EGFR/ErbB2 heterodimers (Spivak-Kroizman et al, 1992;Wada et al, 1990).…”

Serine phosphorylation of paxillin by heregulin-β1: role of p38 mitogen activated protein kinase

“…The HRG-induced increase in serine phosphorylation of paxillin and cell scattering were selectively inhibited by a speci®c inhibitor of p38 MAPK or a dominant-negative p38 MAPK mutant, but not byIntroduction Abnormalities in the expression, structure, or activity of proto-oncogene products contribute to the development and maintenance of the malignant phenotype. For example, HER2 (also known as c-erbB2 or c-neu) encodes a 185 kD transmembrane glycoprotein with intrinsic tyrosine kinase activity (Stern et al, 1986) that has been shown to be overexpressed or ampli®ed, or both, in a number of human malignancies, including breast cancer (Slamon et al, 1987). Overexpression of the HER2 receptor is associated with increased progression and metastasis, an aggressive clinical course, and decreased disease-free survival in human breast cancer patients (Slamon et al, 1989).…”