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Curr Protoc Cell Biol. 2010 Mar;Chapter 15:Unit 15.14. doi: 10.1002/0471143030.cb1514s46.

Quantitative analysis of endocytosis and turnover of epidermal growth factor (EGF) and EGF receptor.

Author information

1
Department of Pharmacology, University of Colorado Denver Medical School, Aurora, Colorado, USA.

Abstract

Binding of epidermal growth factor (EGF) to the EGF receptor (EGFR) initiates signal transduction, ultimately leading to altered gene expression. Ligand-activated EGFR is also rapidly internalized and then targeted to lysosomes for degradation or recycled back to the plasma membrane. Endocytosis is a major regulator of EGFR signaling. Therefore, elucidation of the mechanisms of EGFR endocytosis is essential for a better understanding of EGFR biology. In order to achieve a comprehensive analysis of these mechanisms, reliable methods for measuring the rates of EGFR protein turnover and the rate parameters for individual steps of EGFR endocytic trafficking must be employed. The protocols in this unit describe methodologies to measure the rates of EGFR synthesis and degradation, to monitor EGF-induced down-regulation of surface EGFR, to measure the kinetic rate parameters of internalization, recycling, and degradation of radiolabeled EGF, and to perform radioiodination of EGF by the chloramine T method.

PMID:
20235100
PMCID:
PMC2878126
DOI:
10.1002/0471143030.cb1514s46
[Indexed for MEDLINE]
Free PMC Article

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