Direct involvement of G protein α_q/11 subunit in regulation of muscarinic receptor-mediated sAPPα release

The G_q/11 protein-coupled receptors, such as muscarinic (M1 & M3) receptors, have been shown to regulate the release of a soluble amyloid precursor protein (sAPPaα) produced from α-secretase processing. However, there is no direct evidence for the precise characteristics of G proteins, and the signaling mechanism for the regulation of G _q/11 protein-coupled receptor-mediated sAPPα release is not clearly understood. This study examined whether the muscarinic receptor-mediated release of sAPPα is directly regulated by Gα_q/11 proteins. The HEK293 cells were transiently cotransfected with muscarinic M3 receptors and a dominant-negative minigene construct of the G protein α subunit. The sAPPα release in the media was measured using an antibody specific for sAPP. The sAPPα release enhancement induced by muscarinic receptor stimulation was decreased by a G_q/11 minigene construct, whereas it was not blocked by a control minigene construct (the Gα carboxy peptide in random order, Gα_qR) or Gαi constructs. This indicated a direct role of the Gα_q/11 protein in the regulation of muscarinic M3 receptor-mediated sAPPα release. We also investigated whether the transactivation of the epidermal growth factor receptor (EGFR) by a muscarinic agonist could regulate the sAPPα release in SH-SY5Y cells. Pretreatment of a specific EGFR kinase inhibitor, tyrophostin AG1478 (250 nM), blocked the EGF-stimulated sAPPα release, but did not block the oxoM-stimulated sAPPα release. This demonstrated that the transactivation of the EGFR by muscarinic receptor activation was not involved in the muscarinic receptor-mediated sAPPα release.