Analysis of phospholipase C-γ activation by a truncated receptor tyrosine kinase in vitro

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Abstract

Phospholipase C-γ (PLC-γ) activity increases rapidly upon activation of many receptor tyrosine kinases. To clarify the mechanism of PLC-γ regulation we measured the effect of the intracellular domain of the epidermal growth factor receptor (EGFR-ID), the core tyrosine kinase domain that lacks self-phosphorylation sites (EGFR-KD) and the core self-phosphorylation sites alone (EGFR-CT) on PLC-γ activity in vitro. Tyrosine phosphorylation of PLC-γ was catalyzed by either the cytoplasmic domain or the kinase domain of the EGFR but PLC-γ activity was increased only by the complete cytoplasmic domain of the receptor. Addition of the self-phosphorylation domain alone or in conjunction with the kinase domain did not increase PLC-γ activity. However, both the cytoplasmic domain of the EGFR and the carboxy terminal tail of the EGFR coimmunoprecipitate with PLC-γ. These results suggest that both tyrosine phosphorylation of PLC-γ and binding of PLC-γ to a tyrosine phosphorylated protein are coordinately required for enzyme activation.

Original languageEnglish
Pages (from-to)347-355
Number of pages9
JournalJournal of Biochemistry, Molecular Biology and Biophysics
Volume5
Issue number5
StatePublished - 1 Dec 2001

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Receptor Protein-Tyrosine Kinases
Type C Phospholipases
Chemical activation
Phosphorylation
Tyrosine
Phosphotransferases
In Vitro Techniques
Enzyme Activation
Cytoplasmic and Nuclear Receptors
Epidermal Growth Factor Receptor
Protein-Tyrosine Kinases
Enzymes

Keywords

  • Epidermal growth factor receptor
  • Phospholipase C
  • Tyrosine kinase

Cite this

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abstract = "Phospholipase C-γ (PLC-γ) activity increases rapidly upon activation of many receptor tyrosine kinases. To clarify the mechanism of PLC-γ regulation we measured the effect of the intracellular domain of the epidermal growth factor receptor (EGFR-ID), the core tyrosine kinase domain that lacks self-phosphorylation sites (EGFR-KD) and the core self-phosphorylation sites alone (EGFR-CT) on PLC-γ activity in vitro. Tyrosine phosphorylation of PLC-γ was catalyzed by either the cytoplasmic domain or the kinase domain of the EGFR but PLC-γ activity was increased only by the complete cytoplasmic domain of the receptor. Addition of the self-phosphorylation domain alone or in conjunction with the kinase domain did not increase PLC-γ activity. However, both the cytoplasmic domain of the EGFR and the carboxy terminal tail of the EGFR coimmunoprecipitate with PLC-γ. These results suggest that both tyrosine phosphorylation of PLC-γ and binding of PLC-γ to a tyrosine phosphorylated protein are coordinately required for enzyme activation.",
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AU - Vega, Quinn

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AB - Phospholipase C-γ (PLC-γ) activity increases rapidly upon activation of many receptor tyrosine kinases. To clarify the mechanism of PLC-γ regulation we measured the effect of the intracellular domain of the epidermal growth factor receptor (EGFR-ID), the core tyrosine kinase domain that lacks self-phosphorylation sites (EGFR-KD) and the core self-phosphorylation sites alone (EGFR-CT) on PLC-γ activity in vitro. Tyrosine phosphorylation of PLC-γ was catalyzed by either the cytoplasmic domain or the kinase domain of the EGFR but PLC-γ activity was increased only by the complete cytoplasmic domain of the receptor. Addition of the self-phosphorylation domain alone or in conjunction with the kinase domain did not increase PLC-γ activity. However, both the cytoplasmic domain of the EGFR and the carboxy terminal tail of the EGFR coimmunoprecipitate with PLC-γ. These results suggest that both tyrosine phosphorylation of PLC-γ and binding of PLC-γ to a tyrosine phosphorylated protein are coordinately required for enzyme activation.

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