Journal of Biological Chemistry
Volume 272, Issue 4, 24 January 1997, Pages 2363-2372
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Cell Biology and Metabolism
Identification of Potential Tyrosine-containing Endocytic Motifs in the Carboxyl-tail and Seventh Transmembrane Domain of the Neurokinin 1 Receptor*

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Although agonist-induced endocytosis of G-protein-coupled receptors is critical for receptor desensitization and resensitization, receptor motifs that interact with the endocytic apparatus have not been adequately characterized. We examined the effects of mutating the rat neurokinin-1 receptor on endocytosis using 125I-substance P, fluorescent substance P, and receptor antibodies. Substance P induced rapid internalization of wild-type receptors that were targeted to perinuclear endosomes. Truncation of the C-tail at residues 324, 342, and 354 reduced internalization up to 60% and caused retention of receptors at the cell surface and in superficial endosomes. Mutation of Tyr-341 and Tyr-349 in potential tyrosine-containing endocytic motifs of the C-tail also impaired internalization. A Y305A mutant within the putative NPX2-3Y endocytic motif of the seventh transmembrane domain showed impaired signaling and was minimally expressed at the plasma membrane but was found in cytoplasmic vesicles. In contrast, a Y305F mutant signaled normally and was normally expressed at the plasma membrane but showed impaired internalization. Thus, endocytosis of the neurokinin 1 receptor relies on several tyrosine-containing sequences in the C-tail and seventh transmembrane domain.

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*

This work was supported by National Institutes of Health Grants DK39957, DK43207, and NS21710. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

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Supported by Fellowship Bo1114/1-1 of the Deutsche Forschungsgemeinschaft.