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(Received for publication, April 3, 1996, and in revised form, June 5, 1996)
From the Departments of Proteinase-activated receptor-2 (PAR-2) is a
G-protein-coupled receptor that is expressed by intestinal epithelial
cells, which are episodically exposed to pancreatic trypsin in the
intestinal lumen. Trypsin cleaves PAR-2 to expose a tethered ligand,
which irreversibly activates the receptor. Thus, PAR-2 may desensitize
and resensitize by novel mechanisms. We examined these mechanisms in
kidney epithelial cells, stably expressing human PAR-2, and intestinal
epithelial cells, which naturally express PAR-2. Trypsin stimulated a
prompt increase in [Ca2+]i, due to mobilization
of intracellular Ca2+, followed by a sustained plateau, due
to influx of extracellular Ca2+. Repeated application of
trypsin caused marked desensitization of this response, which is due in
part to (a) irreversible cleavage of the receptor by
trypsin and (b) protein kinase C-mediated termination of
signaling. Trypsin exposure resulted in internalization of PAR-2 into
early endosomes and then lysosomes; but endocytosis was not the
mechanism of rapid desensitization. Thus, activated PAR-2 is
endocytosed and degraded. The Ca2+ response to trypsin
resensitized by 60-90 min. Brefeldin A, which disrupted Golgi stores
of PAR-2, and cycloheximide, which inhibited protein synthesis,
markedly attenuated resensitization. Thus, PAR-2-mediated
Ca2+ mobilization desensitizes by irreversible receptor
cleavage, protein kinase C-mediated termination of signaling, and PAR-2
targeting to lysosomes. It resensitizes by mobilization of large Golgi
stores and synthesis of new receptors.
Volume 271, Number 36,
Issue of September 6, 1996
pp. 22003-22016
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.
,
,
,
Surgery,
¶ Medicine, and 
Physiology, University of
California, San Francisco, California 94143-0660 and the
§ Department of Nutritional Sciences, University of
Calfornia, Berkeley, California 94720
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F. Schmidlin, O. Dery, K. O. DeFea, L. Slice, S. Patierno, C. Sternini, E. F. Grady, and N. W. Bunnett Dynamin and Rab5a-dependent Trafficking and Signaling of the Neurokinin 1 Receptor J. Biol. Chem., June 29, 2001; 276(27): 25427 - 25437. [Abstract] [Full Text] [PDF]
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