Differential regulation of NPR-B/GC-B by protein kinase C and calcium

Abstract

C-type natriuretic peptide (CNP) activation of the guanylyl cyclase-linked natriuretic peptide receptor-B (NPR-B) stimulates vasorelaxation and bone growth. Hormones and phorbol esters (PMA) inhibit NPR-B in calcium-and protein kinase C-dependent manners, respectively. Here, we characterize the kinetic properties of NPR-B in membranes from cells exposed to PMA, the calcium ionophore, ionomycin, or sphingosine-1-phosphate (S1P). PMA and ionomycin primarily increased the K m and decreased the V max of NPR-B for GTP, respectively, whereas S1P caused modest changes in both parameters. PMA and S1P treatment increased the EC 50 for CNP activation by 8 and 3-fold, whereas ionomycin was ineffective. All three agents caused NPR-B dephosphorylation, but the basis for the loss of phosphate differed between treatments. In vitro phosphorylation of NPR-B in membranes was markedly diminished by prior PMA or S1P exposure, whereas ionomycin pretreatment had no effect. The involvement of the known phosphorylated residues in each process was tested with a mutant receptor containing glutamates substituted for these sites. While the effect of PMA was lost on this receptor, the effects of S1P and ionomycin were only partially blocked. Our data suggest that the molecular bases for PMA-and calcium-dependent inhibition of NPR-B are unique. The former results from reduced phosphorylation of a known site and primarily affects the affinity of NPR-B for CNP and GTP. The latter is associated reductions in maximal velocities by a mechanism that does not involve inhibition of NPR-B phosphorylation and requires a process in addition to the dephosphorylation of the known sites.

Dephosphorylation and inactivation of NPR2 guanylyl cyclase in granulosa cells contributes to the LH-induced decrease in cGMP that causes resumption of meiosis in rat oocytes

“…Signaling through various other hormones and growth factors also reduces the guanylyl cyclase activity of NPR2 in other cells, for example, vasopressin (Abbey and Potter, 2002), PDGF (Chrisman and Garbers, 1999;Abbey and Potter, 2003), lysophosphatidic acid (Abbey and Potter, 2003;Potthast et al, 2004), sphingosine-1-phosphate (Abbey-Hosch et al, 2005) and thyrotropin-releasing hormone (Thompson et al, 2014). In studies of sphingosine-1-phosphate acting on cultured fibroblasts that overexpress NPR2 (Abbey-Hosch et al, 2005), there is a correlation between dephosphorylation and the decrease in guanylyl cyclase activity.…”

“…In studies of sphingosine-1-phosphate acting on cultured fibroblasts that overexpress NPR2 (Abbey-Hosch et al, 2005), there is a correlation between dephosphorylation and the decrease in guanylyl cyclase activity. However, LH signaling in the ovarian follicle is the first example of a physiological pathway that is mediated by such a mechanism.…”

The indolocarbazole, Gö6976, inhibits guanylyl cyclase‐A and ‐B

“…Activation of protein kinase C (PKC) by phorbol 12‐myristate, 13‐acetate (PMA) inhibits hormone‐dependent activation of GC‐A and GC‐B in a manner that correlates with site‐specific receptor dephosphorylation (Potter and Garbers, 1994; Potter and Hunter, 2000; Potthast et al ., 2004). Guanylyl cyclase inhibition and dephosphorylation by PMA is blocked by the general PKC inhibitor GF‐109203X, which competes for ATP binding to the active site of PKC (Potter and Garbers, 1994; Abbey‐Hosch et al ., 2005). GF‐109203X has no inhibitory effect on natriuretic peptide receptors in the absence of PKC activators.…”

Selective Regulation of Oocyte Meiotic Events Enhances Progress in Fertility Preservation Methods

“…109 Second, induction of EGF-R activity decreases Npr2 expression in the cumulus cells by means of dephosphorylation reactions. 111 Third, by activating EGF-R, LH increases the secretion of amphiregulin, which leads to downregulation in Nppc expression. 91 Moreover, EGF-R activation inhibits the Nppc mRNA expression in the somatic cells 91 and causes a decline in Nppc concentration.…”