EGF-receptor (EGF-R) tyrosine kinase is required for the down-regulation of activated EGF-R. However, controversy exists as to whether ligand-induced activation of the EGF-R tyrosine kinase is required for internalization or for lysosomal targeting. We have addressed this issue using a cell-free assay that selectively measures the recruitment of EGF-R into coated pits. Here we show that EGF bound to wild-type receptors is efficiently sequestered in coated pits. In contrast, sequestration of kinase-deficient receptors occurs inefficiently and at the same basal rate of endocytosis of unoccupied receptors or receptors lacking any cytoplasmic domain. Sequestration of deletion mutants of the EGF-R that lack autophosphorylation sites also requires an active tyrosine kinase. This suggests that a tyrosine kinase substrate(s) other than the EGF-R itself, is required for its efficient ligand-induced recruitment into coated pits. Addition of a soluble EGF-R tyrosine kinase fully and specifically restores the recruitment of kinase-deficient EGF-R into coated pits providing a powerful functional assay for identification of these substrate(s).
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1 April 1995
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April 01 1995
Recruitment of epidermal growth factor receptors into coated pits requires their activated tyrosine kinase.
C Lamaze,
C Lamaze
Department of Cell Biology, Scripps Research Institute, La Jolla, California 92037.
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S L Schmid
S L Schmid
Department of Cell Biology, Scripps Research Institute, La Jolla, California 92037.
Search for other works by this author on:
C Lamaze
Department of Cell Biology, Scripps Research Institute, La Jolla, California 92037.
S L Schmid
Department of Cell Biology, Scripps Research Institute, La Jolla, California 92037.
Online ISSN: 1540-8140
Print ISSN: 0021-9525
J Cell Biol (1995) 129 (1): 47–54.
Citation
C Lamaze, S L Schmid; Recruitment of epidermal growth factor receptors into coated pits requires their activated tyrosine kinase.. J Cell Biol 1 April 1995; 129 (1): 47–54. doi: https://doi.org/10.1083/jcb.129.1.47
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