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. 1979 Nov;76(11):5689–5693. doi: 10.1073/pnas.76.11.5689

Hormone receptor topology and dynamics: Morphological analysis using ferritin-labeled epidermal growth factor

James A McKanna *, Harry T Haigler †, Stanley Cohen †
PMCID: PMC411715  PMID: 230489

Abstract

Previous studies using a biologically active 1:1 conjugate of EGF and ferritin (F-EGF) have traced the binding and internalization of the hormone molecules. In the present report, we develop ultrastructural criteria for identification of the F-EGF·receptor complex, and, thereby, enable utilization of the F-EGF as an indirect marker to localize the receptor for this peptide hormone. The ferritin cores of bound F-EGF are situated 4-6 nm from the extracellular surface of the membrane. When cells were incubated for up to 30 min at 37°C, this characteristic spatial relationship was observed in all uptake stages (surface clustering, endocytosis, and incorporation into multivesicular bodies), indicating that the hormone·receptor complex remains intact through these steps. However, when incubation was continued for periods sufficient to allow hormone degradation (30-60 min), pools of free ferritin were observed in lysosomes. In the presence of various amine inhibitors of hormone degradation, internalization and multivesicular body incorporation proceeded, but hormone·receptor degradation was blocked as evidenced by preservation of the ferritin—membrane relationship; i.e., no pools of free ferritin were seen after 60 min. These data provide morphological support for the hypothesis that down-regulation of surface receptors involves internalization of intact hormone·receptor complexes. In addition, we have developed a method for viewing the surface of intact cells en face, allowing closer scrutiny of the clustering of F-EGF·receptor complexes in the plane of the membrane prior to internalization. The particles in the F-EGF clusters observed by this method are spaced at 12 nm center-to-center, serving to set upper limits on the packing dimensions of the EGF·receptor complex.

Keywords: endocytosis, lysosomes, lysosome inhibitors, multivesicular bodies

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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