Transmembrane domain sequence requirements for activation of the p185c-neu receptor tyrosine kinase

The receptor tyrosine kinase p185c-neu can be constitutively activated by the transmembrane domain mutation Val664-->Glu, found in the oncogenic mutant p185neu. This mutation is predicted to allow intermolecular hydrogen bonding and receptor dimerization. Understanding the activation of p185c-neu...

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Bibliographic Details
Published in:The Journal of cell biology 1997-05, Vol.137 (3), p.619-631
Main Authors: Chen, L I, Webster, M K, Meyer, A N, Donoghue, D J
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Cell Compartmentation
Cell Membrane - metabolism
Cell Transformation, Neoplastic
Cellular biology
Cloning
Enzyme Activation
Enzymes
Fluorescent Antibody Technique, Indirect
Glutamates - chemistry
Humans
Membrane Proteins - chemistry
Molecular Sequence Data
Mutagenesis, Site-Directed
Mutation
Receptor Protein-Tyrosine Kinases - chemistry
Receptor Protein-Tyrosine Kinases - metabolism
Receptor, ErbB-2 - chemistry
Receptor, ErbB-2 - metabolism
Repetitive Sequences, Nucleic Acid
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Summary:The receptor tyrosine kinase p185c-neu can be constitutively activated by the transmembrane domain mutation Val664-->Glu, found in the oncogenic mutant p185neu. This mutation is predicted to allow intermolecular hydrogen bonding and receptor dimerization. Understanding the activation of p185c-neu has assumed greater relevance with the recent observation that achondroplasia, the most common genetic form of human dwarfism, is caused by a similar transmembrane domain mutation that activates fibroblast growth factor receptor (FGFR) 3. We have isolated novel transforming derivatives of p185c-neu using a large pool of degenerate oligonucleotides encoding variants of the transmembrane domain. Several of the transforming isolates identified were unusual in that they lacked a Glu at residue 664, and others were unique in that they contained multiple Glu residues within the transmembrane domain. The Glu residues in the transforming isolates often exhibited a spacing of seven residues or occurred in positions likely to represent the helical interface. However, the distinction between the sequences of the transforming clones and the nontransforming clones did not suggest clear rules for predicting which specific sequences would result in receptor activation and transformation. To investigate these requirements further, entirely novel transmembrane sequences were constructed based on tandem repeats of simple heptad sequences. Activation was achieved by transmembrane sequences such as [VVVEVVA]n or [VVVEVVV]n, whereas activation was not achieved by a transmembrane domain consisting only of Val residues. In the context of these transmembrane domains, Glu or Gln were equally activating, while Lys, Ser, and Asp were not. Using transmembrane domains with two Glu residues, the spacing between these was systematically varied from two to eight residues, with only the heptad spacing resulting in receptor activation. These results are discussed in the context of activating mutations in the transmembrane domain of FGFR3 that are responsible for the human developmental syndromes achondroplasia and acanthosis nigricans with Crouzon Syndrome.
ISSN:0021-9525
1540-8140
DOI:10.1083/jcb.137.3.619