The erbB-2͞HER2 oncogene is overexpressed in a significant fraction of human carcinomas of the breast, ovary, and lung in a manner that correlates with poor prognosis. Although the encoded protein resembles several receptors for growth factors, no high affinity ligand of ErbB-2 has so far been fully characterized. However, several lines of evidence have raised the possibility that ErbB-2 can augment signal transduction initiated by binding of certain growth factors to their direct receptors. Here, we contrasted these two models of ErbB-2 function: First, examination of a large series of epidermal growth factor (EGF)-like ligands and neuregulins, including virus-encoded ligands as well as related motifs derived from the precursor of EGF, failed to detect interactions with ErbB-2 when this protein was singly expressed. Second, by using antibodies that block inter-ErbB interactions and cells devoid of surface ErbB-2, we learned that signaling by all ligands examined, except those derived from the precursor of EGF, was enhanced by the oncoprotein. These results imply that ErbB-2 evolved as a shared receptor subunit of all ErbB-specific growth factors. Thus, oncogenicity of ErbB-2 in human epithelia may not rely on the existence of a specific ligand but rather on its ability to act as a coreceptor for multiple stroma-derived growth factors.Cellular growth and fate determination are controlled by a large variety of extracellular ligands and specific cell surface receptors. The largest family of such receptors is that of the growth factor receptors with intrinsic tyrosine kinase activity (1). Type-1 tyrosine kinase receptors, also known as ErbB͞ HER proteins, comprise one of the better-characterized subfamilies of growth factor receptors, of which the epidermal growth factor (EGF) receptor (ErbB-1) is the prototype (reviewed in ref.2). The four ErbB members form homo-and heterodimeric complexes on binding of EGF-like or neuregulin (NRG) ligands, and, thereby, their kinase activity is stimulated and intracellular signals are generated. Constitutive stimulation of these pathways through autocrine or other mechanisms is associated with several types of human cancer (3). Most relevant is the frequent overexpression, often as a result of gene amplification, of ErbB-2͞HER2 in breast, ovary, lung, and other types of epithelial cancers (reviewed in refs. 4 and 5). In some tissues, this overexpression was correlated with poorer prognosis and a more aggressive tumor phenotype (6).Although ErbB-2 shares extensive structural homology with other ErbBs both along the catalytic intracellular domain and in the extracellular putative ligand binding region, many attempts to identify stimulatory ligands specific to ErbB-2 have so far failed. For example, detection of an activity that enhances ErbB-2 phosphorylation led to molecular cloning of the Neu differentiation factor (NDF) and heregulin, two of a dozen isoforms of NRG1, all of which bind to ErbB-3 and ErbB-4 (7). Nevertheless, several observations imply that ErbB-2 homodime...

The multiple isoforms of Neu differentiation factor (NDF/neuregulin) induce a pleiotropic cellular response that is isoform-specific and cell type-dependent. The molecular basis of this heterogeneity was addressed by comparing the two major groups of isoforms, ␣ and ␤. Both groups bind to the catalytically impaired receptor tyrosine kinase ErbB-3, whose mitogenic stimulation by NDF requires transactivation by other ErbB proteins, either ErbB-1 or ErbB-2. By expressing each pair of receptors in interleukin 3-dependent myeloid cells, we found that both isoforms induced mitogenic signals in cells co-expressing the combination of ErbB-3 with ErbB-2. However, only the ␤ isoform stimulated cells that expressed both ErbB-3 and ErbB-1, and neither isoform was active on cells expressing ErbB-3 alone. Both isoforms bind to all ErbB-3-expressing cells, albeit with different affinities, but the co-stimulatory mitogenic effect is correlated with the ability of each auxiliary receptor to transphosphorylate ErbB-3. These results imply that NDF isoforms differ in their ability to induce receptor heterodimers; whereas both types of isoforms signal through ErbB-3/ErbB-2 heterodimers, only ␤ isoforms are able to stabilize ErbB-3/ErbB-1 heterodimers.

The primary translation product of barley aspartic proteinase, phytepsin (EC 3.4.23.40), consists of a signal sequence, a propart, and mature enzyme forms. Here, we describe post-translational processing and activation of phytepsin during its transport to the vacuole in roots, as detected by using metabolic labeling and immunoprecipitation. After removal of the signal sequence, the glycosylated precursor of 53 kDa (P53) was produced and further processed to polypeptides of 31 and 15 kDa (P31 ؉ P15) and, subsequently, to polypeptides of 26 and 9 kDa (P26 ؉ P9), 45 min and 24 h after synthesis, respectively. The processing occurred in a late-Golgi compartment or post-Golgi compartment, because brefeldin A inhibited the processing, and P53 acquired partial endoglycosidase H resistance 30 min after synthesis, whereas P15 was completely resistant. The N-glycosylation inhibitor tunicamycin had no effect on transport, but the absence of glycans on P53 accelerated the proteolytic processing. Phytepsin was also expressed in baculovirus-infected insect cells. The recombinant prophytepsin underwent autoproteolytic activation in vitro and showed enzymatic properties similar to the enzyme purified from grains. However, a comparison of the in vitro/in vivo processing sites revealed slight differences, indicating that additional proteases are needed for the completion of the maturation in vivo.