A1 adenosine receptor-stimulated exocytosis in bladder umbrella cells requires phosphorylation of ADAM17 Ser-811 and EGF receptor transactivation.
Bottom Line: Despite the importance of ADAM17-dependent cleavage in normal biology and disease, the physiological cues that trigger its activity, the effector pathways that promote its function, and the mechanisms that control its activity, particularly the role of phosphorylation, remain unresolved.Preventing this phosphorylation event by expression of a nonphosphorylatable ADAM17(S811A) mutant or expression of a tail-minus construct inhibits A1AR-stimulated, ADAM17-dependent HB-EGF cleavage.Furthermore, expression of ADAM17(S811A) in bladder tissues impairs A1AR-induced apical exocytosis.
Affiliation: Departments of Medicine and Cell Biology, University of Pittsburgh, Pittsburgh, PA 15261.Show MeSH
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Mentions: We next addressed how ADAM17 activity is coupled to A1AR activation. Previous studies showed that A1AR signals through Giα to inhibit the activity of adenylyl cyclase, whereas the βγ subunits of Gi increase the activity of phospholipase C-β (PLCβ), which hydrolyzes phosphatidylinositol 4,5-bisphosphate to generate inositol trisphosphate (IP3) and diacylgycerol (Freund et al., 1994; Bucheimer and Linden, 2004; Chang et al., 2008). The latter stimulates the activity of PKC, a well-known regulatory kinase that was previously implicated in ADAM17 activation (Dang et al., 2011; Kveiborg et al., 2011; Lemjabbar-Alaoui et al., 2011). We found that pertussis toxin–mediated inhibition of Gi or inhibition of Gβγ subunit activity by the inhibitor M119K (Kirui et al., 2010) significantly impaired CCPA-mediated apical exocytosis in rabbit bladder umbrella cells (Figure 5A). Furthermore, the PLC-selective antagonist U73122 caused marked inhibition of CCPA-induced changes in CT (Figure 5A). Thus ADAM17 activation downstream of A1AR likely occurs by way of a classical Gi-stimulated signaling cascade involving Gβγ and PLCβ.
Affiliation: Departments of Medicine and Cell Biology, University of Pittsburgh, Pittsburgh, PA 15261.