Trafficking of proteinase-activated receptor-2 and beta-arrestin-1 tagged with green fluorescent protein. beta-Arrestin-dependent endocytosis of a proteinase receptor.
Proteases cleave proteinase-activated receptors (PARs) to expose N-terminal tethered ligands that bind and activate the cleaved receptors. The tethered ligand, once exposed, is always available to interact with its binding site. Thus, efficient mechanisms must prevent continuous activation, including receptor phosphorylation and uncoupling from G-proteins, receptor endocytosis, and lysosomal degradation. beta-Arrestins mediate uncoupling and endocytosis of certain neurotransmitter receptors, which are activated in a reversible manner. However, the role of beta-arrestins in trafficking of PARs, which are irreversibly activated, and the effects of proteases on the subcellular distribution of beta-arrestins have not been examined. We studied trafficking of PAR2 and beta-arrestin1 coupled to green fluorescent protein. Trypsin induced the following: (a) redistribution of beta-arrestin1 from the cytosol to the plasma membrane, where it co-localized with PAR2; (b) internalization of beta-arrestin1 and PAR2 into the same early endosomes; (c) redistribution of beta-arrestin1 to the cytosol concurrent with PAR2 translocation to lysosomes; and (d) mobilization of PAR2 from the Golgi apparatus to the plasma membrane. Overexpression of a C-terminal fragment of beta-arrestin-319-418, which interacts constitutively with clathrin but does not bind receptors, inhibited agonist-induced endocytosis of PAR2. Our results show that beta-arrestins mediate endocytosis of PAR2 and support a role for beta-arrestins in uncoupling of PARs.
Déry
O
O
Department of Surgery, University of California, San Francisco, California 94143-0660, USA.
Thoma
M S
MS
Wong
H
H
Grady
E F
EF
Bunnett
N W
NW
D000818:Animals; D019390:Arrestins / Q000378:metabolism*; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D004705:Endocytosis; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D011817:Rabbits; D051381:Rats; D044464:Receptor, PAR-2; D018179:Receptors, Thrombin / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism; D014407:Tumor Cells, Cultured; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins
C000605256:Arrb1 protein, rat; D019390:Arrestins; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D044464:Receptor, PAR-2; D018179:Receptors, Thrombin; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins; D049452:Green Fluorescent Proteins; D014357:Trypsin; D002118:Calcium