Activation of Neu (ErbB-2) mediated by disulfide bond-induced dimerization reveals a receptor tyrosine kinase dimer interface.
Receptor dimerization is a crucial intermediate step in activation of signaling by receptor tyrosine kinases (RTKs). However, dimerization of the RTK Neu (also designated ErbB-2, HER-2, and p185(neu)), while necessary, is not sufficient for signaling. Earlier work in our laboratory had shown that introduction of an ectopic cysteine into the Neu juxtamembrane domain induces Neu dimerization but not signaling. Since Neu signaling does require dimerization, we hypothesized that there are additional constraints that govern signaling ability. With the importance of the interreceptor cross-phosphorylation reaction, a likely constraint was the relative geometry of receptors within the dimer. We have tested this possibility by constructing a consecutive series of cysteine substitutions in the Neu juxtamembrane domain in order to force dimerization along a series of interreceptor faces. Within the group that dimerized constitutively, a subset had transforming activity. The substitutions in this subset all mapped to the same face of a predicted alpha helix, the most likely conformation for the intramembrane domain. Furthermore, this face of interaction aligns with the projected Neu* V664E substitution and with a predicted amphipathic interface in the Neu juxtamembrane domain. We propose that these results identify an RTK dimer interface and that dimerization of this RTK induces an extended contact between juxtamembrane and intramembrane alpha helices.
Department of Pathology, Yale University, New Haven, Connecticut 06520-8023, USA.
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D016475:3T3 Cells; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D003545:Cysteine; D017931:DNA Primers; D019281:Dimerization; D004220:Disulfides; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D017433:Protein Structure, Secondary; D051381:Rats; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment
D003545:Cysteine; D017931:DNA Primers; D004220:Disulfides; D020794:Receptor Protein-Tyrosine Kinases; D018719:Erb-b2 Receptor Tyrosine Kinases; D011994:Recombinant Proteins