The role of the C-terminal tail in protease-activated receptor-2-mediated Ca 2+ signalling, proline-rich tyrosine kinase-2 activation, and mitogen-activated protein kinase activity

Abstract

C-terminal truncation mutants were made to investigate the role of the C-terminus in coupling proteinase-activated receptor-2 (PAR-2) to various signalling pathways. Membrane expression of the δ15, δ34, δ43, and δ34–43 mutants was similar; however, expression of δtail was lost, as was agonist-mediated internalisation of δtail, δ43, and δ34–43. Additionally, trypsin and SLIGKV-stimulated [ 3H]IP accumulation was abrogated in cells transiently expressing δ43 or δ34–43 truncations, but remained unaffected in cells expressing δ34 or δ15. PAR-2 agonist-stimulated intracellular Ca 2+ mobilisation and PYK-2 activity were also abolished by δtail, δ43, and δ34–43 mutants. However, trypsin-stimulated stress-activated protein kinases (SAPKs) or extracellular signal-regulated kinase (ERK) activities were unaffected by the δ34–43 mutation, although activity was abrogated following δ43 or δtail truncations, suggesting that Ca 2+ mobilisation, PYK-2, or receptor internalisation are not requied for activation of SAPKs or ERK. These studies identify a novel sequence within the PAR-2 C-terminus essential for InsP 3 generation and PYK-2 activity but not mitogen-activated protein kinase (MAPK) activation.