Transactivation of the PAR1-PAR2 heterodimer by thrombin elicits β-arrestin-mediated endosomal signaling

J Biol Chem. 2013 Apr 19;288(16):11203-15. doi: 10.1074/jbc.M112.439950. Epub 2013 Mar 8.

Abstract

Thrombin cleaves the N terminus of PAR1, generating a new N-terminal domain that functions as a tethered ligand that binds intermolecularly to activate PAR2 in trans. The mechanisms that regulate PAR1-PAR2 heterodimer signaling and trafficking are not known. We now report that PAR1 and PAR2 form a heterodimer that exhibits unique trafficking and signaling behaviors compared with receptor protomers. Using bioluminescence resonance energy transfer, immunofluorescence microscopy, co-immunoprecipitation, and cells expressing receptors exogenously and endogenously, we show that PAR1 and PAR2 specifically interact and form stable dimers. Intriguingly, the PAR1-PAR2 heterodimer displays constitutive internalization that is driven by PAR1 C-terminal tail sorting motifs and is a process that enhances dimer formation. Upon thrombin activation, PAR1-PAR2 dimers co-internalize and recruit β-arrestins to endosomes. Remarkably, PAR1-PAR2 heterodimers appear to utilize a distinct interface for β-arrestin interaction compared with receptor protomers. Moreover, thrombin-activated PAR1-PAR2 heterodimers enhance β-arrestin-mediated ERK1/2 activation in the cytoplasm, whereas activated ERK1/2 induced by the thrombin-activated PAR1 protomer redistributes to the nucleus. Thus, the formation of PAR1-PAR2 heterodimers provides additional modes of thrombin-stimulated signaling responses that appear to be distinctly regulated compared with the receptor protomer.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Motifs
  • Animals
  • Arrestins / genetics
  • Arrestins / metabolism*
  • COS Cells
  • Chlorocebus aethiops
  • Endosomes / genetics
  • Endosomes / metabolism*
  • HeLa Cells
  • Humans
  • Promoter Regions, Genetic / physiology
  • Protein Multimerization / physiology*
  • Receptor, PAR-1 / genetics
  • Receptor, PAR-1 / metabolism*
  • Receptor, PAR-2 / genetics
  • Receptor, PAR-2 / metabolism*
  • Signal Transduction / physiology*
  • Thrombin / genetics
  • Thrombin / metabolism*
  • Transcriptional Activation / physiology*
  • beta-Arrestins

Substances

  • Arrestins
  • Receptor, PAR-1
  • Receptor, PAR-2
  • beta-Arrestins
  • Thrombin