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

Chen LI, Webster MK, Meyer AN, Donoghue DJ - J. Cell Biol. (1997)

Bottom Line: The receptor tyrosine kinase p185c-neu can be constitutively activated by the transmembrane domain mutation Val664-->Glu, found in the oncogenic mutant p185neu.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry and Center for Molecular Genetics, University of California, San Diego, La Jolla 92093-0367, USA.

ABSTRACT
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.

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Oncoproteins and receptors activated by mutations in  the transmembrane domain. Amino acids are shown that allow  activation of p185c-neu when substituted at residue 664, activation  of BPV-E5 when substituted at residue 17, and activation of  FGFR3 when substituted at residue 380, as discussed in the text.  Those substitutions that allow activation in these three different  systems share the property that they are strongly polar in an otherwise hydrophobic membrane environment, and thus share the  ability to participate in hydrogen bond formation that may stabilize dimer formation.
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Figure 10: Oncoproteins and receptors activated by mutations in the transmembrane domain. Amino acids are shown that allow activation of p185c-neu when substituted at residue 664, activation of BPV-E5 when substituted at residue 17, and activation of FGFR3 when substituted at residue 380, as discussed in the text. Those substitutions that allow activation in these three different systems share the property that they are strongly polar in an otherwise hydrophobic membrane environment, and thus share the ability to participate in hydrogen bond formation that may stabilize dimer formation.

Mentions: As summarized in Fig. 10, prior studies have demonstrated that substitutions at position 664 of p185neu exhibit the following pattern of biological activity: Glu, Gln > Asp, Tyr ≫ Val, Lys, Gly, His (Bargmann and Weinberg, 1988b). These results are generally consistent with a role of polar or hydrophilic residues in promoting activation, although the failure of Lys and His to allow for activation remains to be explained. The E5 oncoprotein of bovine papilloma virus (BPV) provides another example of the importance of the transmembrane domain for activation, since this 44residue integral membrane protein requires dimerization and disulfide bond formation for its biological activity (Horwitz et al., 1988, 1989; Goldstein et al., 1992). A key requirement of its hydrophobic transmembrane domain is the presence of Gln17, which appears to participate in interhelical hydrogen bonding and may be substituted by either Glu or Lys, and to a lesser extent by His (Meyer et al., 1994). Significantly, the transmembrane domain of E5 can be largely replaced by the membrane-spanning region from activated p185neu (Meyer et al., 1994).


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

Chen LI, Webster MK, Meyer AN, Donoghue DJ - J. Cell Biol. (1997)

Oncoproteins and receptors activated by mutations in  the transmembrane domain. Amino acids are shown that allow  activation of p185c-neu when substituted at residue 664, activation  of BPV-E5 when substituted at residue 17, and activation of  FGFR3 when substituted at residue 380, as discussed in the text.  Those substitutions that allow activation in these three different  systems share the property that they are strongly polar in an otherwise hydrophobic membrane environment, and thus share the  ability to participate in hydrogen bond formation that may stabilize dimer formation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139875&req=5

Figure 10: Oncoproteins and receptors activated by mutations in the transmembrane domain. Amino acids are shown that allow activation of p185c-neu when substituted at residue 664, activation of BPV-E5 when substituted at residue 17, and activation of FGFR3 when substituted at residue 380, as discussed in the text. Those substitutions that allow activation in these three different systems share the property that they are strongly polar in an otherwise hydrophobic membrane environment, and thus share the ability to participate in hydrogen bond formation that may stabilize dimer formation.
Mentions: As summarized in Fig. 10, prior studies have demonstrated that substitutions at position 664 of p185neu exhibit the following pattern of biological activity: Glu, Gln > Asp, Tyr ≫ Val, Lys, Gly, His (Bargmann and Weinberg, 1988b). These results are generally consistent with a role of polar or hydrophilic residues in promoting activation, although the failure of Lys and His to allow for activation remains to be explained. The E5 oncoprotein of bovine papilloma virus (BPV) provides another example of the importance of the transmembrane domain for activation, since this 44residue integral membrane protein requires dimerization and disulfide bond formation for its biological activity (Horwitz et al., 1988, 1989; Goldstein et al., 1992). A key requirement of its hydrophobic transmembrane domain is the presence of Gln17, which appears to participate in interhelical hydrogen bonding and may be substituted by either Glu or Lys, and to a lesser extent by His (Meyer et al., 1994). Significantly, the transmembrane domain of E5 can be largely replaced by the membrane-spanning region from activated p185neu (Meyer et al., 1994).

Bottom Line: The receptor tyrosine kinase p185c-neu can be constitutively activated by the transmembrane domain mutation Val664-->Glu, found in the oncogenic mutant p185neu.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry and Center for Molecular Genetics, University of California, San Diego, La Jolla 92093-0367, USA.

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus