Cell Biology and Metabolism
The N-terminal Domain of a Glycolipid-anchored Prion Protein Is Essential for Its Endocytosis via Clathrin-coated Pits

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The cellular prion protein (PrPC) is a glycolipid-anchored protein that is involved in the pathogenesis of fatal spongiform encephalopathies. We have shown previously that, in contrast to several other glycolipid-anchored proteins, chPrP, the chicken homologue of mammalian PrPC, is endocytosed via clathrin-coated pits in cultured neuroblastoma cells, as well as in embryonic neurons and glia (Shyng, S.-L., Heuser, J. E., and Harris, D. A.(1994) J. Cell Biol. 125, 1239-1250). In this study, we have determined that the N-terminal half of the chPrP polypeptide chain is essential for its endocytosis. Deletions within this region reduce the amount of chPrP internalized, as measured by surface iodination or biotinylation, and decrease its concentration in clathrin-coated pits, as determined by quantitative electron microscopic immunogold labeling. Mouse PrP, as well as two mouse PrP/chPrP chimeras, are internalized as efficiently as chPrP, suggesting that conserved features of secondary and tertiary structure are involved in interaction with the endocytic machinery. Our results indicate that the ectodomain of a protein can contain endocytic targeting information, and they strongly support a model in which the polypeptide chain of PrPC binds to the extracellular domain of a transmembrane protein that contains a coated pit localization signal in its cytoplasmic tail.

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This work was supported in part by grants (to D. A. H.) from the National Institutes of Health (Grant NS30137), the Alzheimer's Association (Richard F. Bristor Investigator-Initiated Research Grant), the March of Dimes Birth Defects Foundation (Basic Research Grant 0571), and the McDonnell Center for Cellular and Molecular Neurobiology at Washington University. Portions of this paper have been presented in abstract form (Harris et al., 1993a). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

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Recipient of fellowship support from the McDonnell Center for Cellular and Molecular Neurobiology, and from National Institutes of Health Postdoctoral Training Grant T32 NS07071.