Separate Signals for Agonist-independent and Agonist-triggered Trafficking of Protease-activated Receptor 1*

Abstract

Protease-activated receptor 1 (PAR1), a G protein-coupled, protease-activated receptor for the serine protease thrombin, is activated when thrombin cleaves its amino-terminal exodomain. This irreversible mechanism of activation may have necessitated an unusual pattern of receptor trafficking. Unactivated PAR1 cycles tonically between the cell surface and an intracellular pool, providing an intracellular store of uncleaved receptors and allowing repopulation of the surface with uncleaved receptors after thrombin exposure without new receptor synthesis. Activated PAR1 internalizes rapidly and is degraded in lysosomes. We report characterization of a PAR1 mutant that trafficked like the wild-type receptor when activated but did not internalize and recycle in the absence of agonist. This complements a previous study in which a mutant with normal tonic internalization but defective agonist-triggered internalization was described. These observations suggest that the trafficking behaviors of unactivated and activated PAR1 are specified by distinct signals within the receptor and imply that PAR1 internalization in the presence or absence of agonist may be mediated by distinct molecular machinery. PAR1 mutants that did not internalize in the absence of agonist were also shown to localize exclusively to the cell surface and to be defective in their ability to repopulate the cell surface with uncleaved receptors after thrombin exposure. These observations suggest that tonic internalization is necessary for maintenance of the intracellular PAR1 pool.

Footnotes

  • * This work was supported by National Institutes of Health Grant HL44907 and by a postdoctoral fellowship from the American Cancer Society (to M. J. S.).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.

  • To whom correspondence should be addressed: Cardiovascular Research Institute, University of California, San Francisco, HSE-1300, 505 Parnassus Ave., San Francisco, CA 94143-0130. Tel.: 415-476-6174; Fax: 415-476-8173; E-mail: shaun_coughlin{at}quickmail.ucsf.edu.

  • Abbreviations:
    PAR
    protease-activated receptor
    PBS
    phosphate-buffered saline
    WT
    wild-type PAR1.
    • Received June 12, 1998.
    • Revision received August 7, 1998.
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