Modulation of erbB kinase activity and oncogenic potential by single point mutations in the glycine loop of the catalytic domain

Hui Kuo G. Shu, Chi Ming Chang, Lakshmeswari Ravi, Lei Ling, Chris M. Castellano, Elizabeth Walter, Robert J. Pelley, Hsing Jien Kung

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Avian c-erbB is activated to a leukemia oncogene following truncation of its amino-terminal ligand-binding domain by retroviral insertion. The insertionally activated transcripts encode protein products which have constitutive tyrosine kinase activity and can induce erythroleukemia but not sarcomas. We have previously found that a valine-to-isoleucine point mutation at position 157 (V157I mutant) within the tyrosine kinase domain of this truncated erbB can dramatically activate the sarcomagenic potential of the oncogene and increase the kinase activity of this oncoprotein. This mutation lies at position 157 of the insertionally activated c-erbB product, affecting a highly conserved valine residue of the glycine loop involved in ATP binding and phosphate transfer. To investigate the functional importance of this residue in the catalytic activity of kinases, we have introduced at this position, by site-directed mutagenesis, codons representing the remaining 18 amino acid residues. Most of the mutants have diminished activity, with six of them completely devoid of kinase activity, indicating the sensitivity of this region to conformational changes. Some of these mutants displayed increased kinase activity and greater transforming potential in comparison with 1A c-erbB, but none had levels as high as those of the V157I mutant. In general, the sarcomagenic potential of the various erbB mutants correlated with their autophosphorylation state and their ability to cause phosphorylation of MAP kinase. However, there are important exceptions such as the V157G mutant, which lacks enhanced autophosphorylation but is highly sarcomagenic. Studies of this and other autophosphorylation site mutants point to the existence of an autophosphorylation-independent pathway in sarcomagenesis. The requirement for leukemogenic potential is much less stringent and correlates with positivity of kinase activity. When the valine- to-isoleucine substitution was put in context of the full-length erbB protein, the mutation relaxed the ligand dependence and had a positive effect on the transforming potential of the full-length c-erbB.

Original languageEnglish
Pages (from-to)6868-6878
Number of pages11
JournalMolecular and Cellular Biology
Volume14
Issue number10
DOIs
Publication statusPublished - Jan 1 1994
Externally publishedYes

Fingerprint

Point Mutation
Glycine
Catalytic Domain
Phosphotransferases
Valine
Isoleucine
Oncogenes
Protein-Tyrosine Kinases
Ligands
Leukemia, Erythroblastic, Acute
Mutation
Oncogene Proteins
Site-Directed Mutagenesis
Codon
Sarcoma
Leukemia
Proteins
Adenosine Triphosphate
Phosphates
Phosphorylation

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

Cite this

Modulation of erbB kinase activity and oncogenic potential by single point mutations in the glycine loop of the catalytic domain. / Shu, Hui Kuo G.; Chang, Chi Ming; Ravi, Lakshmeswari; Ling, Lei; Castellano, Chris M.; Walter, Elizabeth; Pelley, Robert J.; Kung, Hsing Jien.

In: Molecular and Cellular Biology, Vol. 14, No. 10, 01.01.1994, p. 6868-6878.

Research output: Contribution to journalArticle

Shu, Hui Kuo G. ; Chang, Chi Ming ; Ravi, Lakshmeswari ; Ling, Lei ; Castellano, Chris M. ; Walter, Elizabeth ; Pelley, Robert J. ; Kung, Hsing Jien. / Modulation of erbB kinase activity and oncogenic potential by single point mutations in the glycine loop of the catalytic domain. In: Molecular and Cellular Biology. 1994 ; Vol. 14, No. 10. pp. 6868-6878.
@article{721188a3cfeb44809ce4a4cc592a2eb6,
title = "Modulation of erbB kinase activity and oncogenic potential by single point mutations in the glycine loop of the catalytic domain",
abstract = "Avian c-erbB is activated to a leukemia oncogene following truncation of its amino-terminal ligand-binding domain by retroviral insertion. The insertionally activated transcripts encode protein products which have constitutive tyrosine kinase activity and can induce erythroleukemia but not sarcomas. We have previously found that a valine-to-isoleucine point mutation at position 157 (V157I mutant) within the tyrosine kinase domain of this truncated erbB can dramatically activate the sarcomagenic potential of the oncogene and increase the kinase activity of this oncoprotein. This mutation lies at position 157 of the insertionally activated c-erbB product, affecting a highly conserved valine residue of the glycine loop involved in ATP binding and phosphate transfer. To investigate the functional importance of this residue in the catalytic activity of kinases, we have introduced at this position, by site-directed mutagenesis, codons representing the remaining 18 amino acid residues. Most of the mutants have diminished activity, with six of them completely devoid of kinase activity, indicating the sensitivity of this region to conformational changes. Some of these mutants displayed increased kinase activity and greater transforming potential in comparison with 1A c-erbB, but none had levels as high as those of the V157I mutant. In general, the sarcomagenic potential of the various erbB mutants correlated with their autophosphorylation state and their ability to cause phosphorylation of MAP kinase. However, there are important exceptions such as the V157G mutant, which lacks enhanced autophosphorylation but is highly sarcomagenic. Studies of this and other autophosphorylation site mutants point to the existence of an autophosphorylation-independent pathway in sarcomagenesis. The requirement for leukemogenic potential is much less stringent and correlates with positivity of kinase activity. When the valine- to-isoleucine substitution was put in context of the full-length erbB protein, the mutation relaxed the ligand dependence and had a positive effect on the transforming potential of the full-length c-erbB.",
author = "Shu, {Hui Kuo G.} and Chang, {Chi Ming} and Lakshmeswari Ravi and Lei Ling and Castellano, {Chris M.} and Elizabeth Walter and Pelley, {Robert J.} and Kung, {Hsing Jien}",
year = "1994",
month = "1",
day = "1",
doi = "10.1128/MCB.14.10.6868",
language = "English",
volume = "14",
pages = "6868--6878",
journal = "Molecular and Cellular Biology",
issn = "0270-7306",
publisher = "American Society for Microbiology",
number = "10",

}

TY - JOUR

T1 - Modulation of erbB kinase activity and oncogenic potential by single point mutations in the glycine loop of the catalytic domain

AU - Shu, Hui Kuo G.

AU - Chang, Chi Ming

AU - Ravi, Lakshmeswari

AU - Ling, Lei

AU - Castellano, Chris M.

AU - Walter, Elizabeth

AU - Pelley, Robert J.

AU - Kung, Hsing Jien

PY - 1994/1/1

Y1 - 1994/1/1

N2 - Avian c-erbB is activated to a leukemia oncogene following truncation of its amino-terminal ligand-binding domain by retroviral insertion. The insertionally activated transcripts encode protein products which have constitutive tyrosine kinase activity and can induce erythroleukemia but not sarcomas. We have previously found that a valine-to-isoleucine point mutation at position 157 (V157I mutant) within the tyrosine kinase domain of this truncated erbB can dramatically activate the sarcomagenic potential of the oncogene and increase the kinase activity of this oncoprotein. This mutation lies at position 157 of the insertionally activated c-erbB product, affecting a highly conserved valine residue of the glycine loop involved in ATP binding and phosphate transfer. To investigate the functional importance of this residue in the catalytic activity of kinases, we have introduced at this position, by site-directed mutagenesis, codons representing the remaining 18 amino acid residues. Most of the mutants have diminished activity, with six of them completely devoid of kinase activity, indicating the sensitivity of this region to conformational changes. Some of these mutants displayed increased kinase activity and greater transforming potential in comparison with 1A c-erbB, but none had levels as high as those of the V157I mutant. In general, the sarcomagenic potential of the various erbB mutants correlated with their autophosphorylation state and their ability to cause phosphorylation of MAP kinase. However, there are important exceptions such as the V157G mutant, which lacks enhanced autophosphorylation but is highly sarcomagenic. Studies of this and other autophosphorylation site mutants point to the existence of an autophosphorylation-independent pathway in sarcomagenesis. The requirement for leukemogenic potential is much less stringent and correlates with positivity of kinase activity. When the valine- to-isoleucine substitution was put in context of the full-length erbB protein, the mutation relaxed the ligand dependence and had a positive effect on the transforming potential of the full-length c-erbB.

AB - Avian c-erbB is activated to a leukemia oncogene following truncation of its amino-terminal ligand-binding domain by retroviral insertion. The insertionally activated transcripts encode protein products which have constitutive tyrosine kinase activity and can induce erythroleukemia but not sarcomas. We have previously found that a valine-to-isoleucine point mutation at position 157 (V157I mutant) within the tyrosine kinase domain of this truncated erbB can dramatically activate the sarcomagenic potential of the oncogene and increase the kinase activity of this oncoprotein. This mutation lies at position 157 of the insertionally activated c-erbB product, affecting a highly conserved valine residue of the glycine loop involved in ATP binding and phosphate transfer. To investigate the functional importance of this residue in the catalytic activity of kinases, we have introduced at this position, by site-directed mutagenesis, codons representing the remaining 18 amino acid residues. Most of the mutants have diminished activity, with six of them completely devoid of kinase activity, indicating the sensitivity of this region to conformational changes. Some of these mutants displayed increased kinase activity and greater transforming potential in comparison with 1A c-erbB, but none had levels as high as those of the V157I mutant. In general, the sarcomagenic potential of the various erbB mutants correlated with their autophosphorylation state and their ability to cause phosphorylation of MAP kinase. However, there are important exceptions such as the V157G mutant, which lacks enhanced autophosphorylation but is highly sarcomagenic. Studies of this and other autophosphorylation site mutants point to the existence of an autophosphorylation-independent pathway in sarcomagenesis. The requirement for leukemogenic potential is much less stringent and correlates with positivity of kinase activity. When the valine- to-isoleucine substitution was put in context of the full-length erbB protein, the mutation relaxed the ligand dependence and had a positive effect on the transforming potential of the full-length c-erbB.

UR - http://www.scopus.com/inward/record.url?scp=0028108969&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0028108969&partnerID=8YFLogxK

U2 - 10.1128/MCB.14.10.6868

DO - 10.1128/MCB.14.10.6868

M3 - Article

VL - 14

SP - 6868

EP - 6878

JO - Molecular and Cellular Biology

JF - Molecular and Cellular Biology

SN - 0270-7306

IS - 10

ER -