Dual bradykinin B2 receptor signalling in A431 human epidermoid carcinoma cells: activation of protein kinase C is counteracted by a GS-mediated stimulation of the cyclic AMP pathway

Abstract

Cell membranes of the human epidermoid cell line A431 express classical bradykinin (BK) B2 receptors, as assessed by [3H]BK binding studies. Furthermore, stimulation by BK induced a time-dependent modulation of protein kinase C (PKC) activity in A431 cells: a rapid activation (t1/2 approximately 1 min) is followed by a slow inhibition (t1/2 approximately 20 min) of PKC translocation measured by [3H]phorbol 12,13-dibutyrate binding. In addition, BK stimulated both adenylate cyclase activity in A431 membranes and accumulation of intracellular cyclic AMP (cAMP) in intact cells in a retarded manner. A possible BK-induced activation of the cAMP pathway mediated via PKC, phospholipase D, prostaglandins or Ca2+/calmodulin was excluded. A 35 kDa protein was found in A431 membranes to be specifically phosphorylated in the presence of both BK and protein kinase A (PKA). An anti-alpha s-antibody, AS 348, abolished stimulation of adenylate cyclase activity in response to BK, cholera toxin and isoprenaline, strongly suggesting the involvement of Gs proteins in the BK action. The BK-activated cAMP signalling system might be important for the observed inactivation of PKC slowly evoked by BK: the BK-induced rapid activation of PKC is decreased by dibutyryl cAMP, and the slow inhibition of PKC is prevented by an inhibitor of PKA, adenosine 3':5'-monophosphothioate (cyclic, Rp isomer). The inhibition of PKC translocation might be exerted directly at the level of PKC activation, since stimulation of phosphoinositide hydrolysis by BK was affected by neither dibutyryl cAMP nor forskolin. Thus our results provide the first evidence that A431 cells BK is able to activate two independent signal-transduction pathways via a single class of B2 receptors but two different G proteins. The lagging stimulation of the cAMP signalling pathway via Gs might serve to switch off PKC, which is rapidly activated via Gq-mediated stimulation of phosphoinositide hydrolysis.

Facilitation of sympathetic neurotransmission by phosphatidylinositol-4,5-bisphosphate-dependent regulation of KCNQ channels in rat mesenteric arteries

“…3 The B 2 receptors are G-protein-coupled receptors and are mainly described as coupled with G q/11 , 40,41 although this receptor interacts with other G proteins, namely G s and G i . [42][43][44] In the present study, the blockade of phospholipase C, which is activated by G q/11 , substantially diminished the action of bradykinin. Meanwhile, pretreatment with H-89 or AACOCF 3 to inhibit the G s -cAMP-protein kinase A pathway or G i -dependent phospholipase A2 activation, respectively, did not alter the effect of bradykinin.…”

A Novel Mitogenic Signaling Pathway of Bradykinin in the Human Colon Carcinoma Cell Line SW-480 Involves Sequential Activation of a Gq/11 Protein, Phosphatidylinositol 3-Kinase β, and Protein Kinase Cε

“…Among the PTX-insensitive G proteins expressed in SW-480 cells, G 12/13 do not stimulate phosphatidylinositol hydrolysis (36) and may be excluded from linking the bradykinin receptor to phospholipase C␤. The bradykinin receptor appears to be capable of interacting with multiple G proteins, including also G s (23,37). If the effect of bradykinin on MAPK is triggered by ␤␥-complexes released from a G s protein as demonstrated for the ␤-adrenergic receptor (9), it might be expected that permanent activation of G s in the presence of CTX simulates or potentiates the BK action on MAPK.…”

“…Phosphatidylinositol Turnover-Determination of total inositol phosphates was performed as described previously (23). In brief, SW-480 cells in 24-well plates were prelabeled with 4 Ci/ml myo- [ 3 H]inositol for 24 h. At 2 h prior to stimulation, the cells were incubated in serum-free medium containing 20 mM HEPES, pH 7.4, and 1 M captopril.…”

Bradykinin promotes neuron-generating division of neural progenitor cells through ERK activation

“…2A; P=0.012). The B2BkR can also catalyze the exchange of GTP for GDP on the α subunits of G i and G s proteins (Blaukat et al, 2000;Hall et al, 1993;Hanke et al, 2006;Liebmann et al, 1996Liebmann et al, , 1990, even though it has been often described as a prototypical G q -protein-coupled receptor. Thus, we assessed if the B2BkR could also be coupled to any member of the G s or G i protein family in NPCs or if the G q protein and its downstream pathway could affect bradykinin-stimulated cAMP production.…”