We have recently reported that two typical G s -coupled receptors, the ␤ 2 -adrenergic receptor and the receptor for prostaglandin E 1 , stimulate phospholipase C-⑀ (PLC- Activation of mitogen-activated protein (MAP) 1 kinases plays a prominent role in many cellular responses to a large variety of membrane receptors. Initially identified as signal transducers of growth factor receptors with intrinsic tyrosine kinase activity, activation of MAP kinases, specifically of the extracellular signal-regulated kinases 1 and 2 (ERK1/2), is now also recognized as a major signal transduction pathway of many receptors coupled to heterotrimeric G proteins. ERK activation by these second messenger-generating receptors is apparently accomplished by diverse molecular mechanisms, depending on the cell types studied as well as the receptor and the heterotrimeric G protein involved (1-5).⑀The cAMP-producing G s -coupled receptors play a rather unique role in ERK activation. These receptors inhibit ERK activation by growth factor receptors in several cell types while stimulating this cellular response in others, most notably in neuronal and endocrine cells. Several models have been proposed to explain these diverse actions of cAMP and G s -coupled receptors on ERK activation, both apparently involving in most cases the principal cAMP target, the cAMP-activated protein kinase (PKA) (6, 7). In particular, PKA-dependent ERK activation in HEK-293 cells by the ␤ 2 -adrenergic receptor (␤ 2 -AR), a prototypical G s -coupled receptor, has been extensively studied. Lefkowitz and coworkers (8) reported that cAMP-activated PKA phosphorylates the ␤ 2 -AR and thereby alters the coupling specificity of the receptor from G s to G i proteins. By this G protein "switching," the ␤ 2 -AR then apparently induces ERK activation by releasing G␤␥ dimers from the pertussis toxin (PTX)-sensitive G i proteins, followed by activation of the cytosolic tyrosine kinase c-Src, the GTPase Ras, and the MAP kinase kinase kinase, Raf-1. Meanwhile, it has been reported that the switching of ␤ 2 -AR from G s to G i proteins is controlled by ␤-arrestin, which recruits the cAMP-degrading phosphodiesterase 4 to the plasma membrane and thereby alters the activity state of PKA (9). In contrast, Schmitt and Stork (10) report that ERK activation by the ␤ 2 -AR in HEK-293 cells, although also PKA-and c-Src-dependent, is PTX-insensitive and thus apparently does not require ␤ 2 -AR switching to G i proteins, a finding recently confirmed by others using a PKAinsensitive ␤ 2 -AR mutant (11). Furthermore, Schmitt and Stork (10) show that ERK activation by ␤ 2 -AR is not mediated by Ras but by the related GTPase Rap1, activating the MAP kinase kinase kinase B-Raf, but not Raf-1. Interestingly, activation of the ␤ 2 -AR also resulted in Ras activation, but this receptor action was apparently independent of cAMP and PKA (10).

Receptor tyrosine kinase regulation of phospholipase C-ε (PLC-ε), which is under the control of Ras-like and Rho GTPases, was studied with HEK-293 cells endogenously expressing PLC-coupled epidermal growth factor (EGF) receptors. PLC and Ca2+ signaling by the EGF receptor, which activated both PLC-γ1 and PLC-ε, was specifically suppressed by inactivation of Ras-related GTPases with clostridial toxins and expression of dominant-negative Rap2B. EGF induced rapid and sustained GTP loading of Rap2B, binding of Rap2B to PLC-ε, and Rap2B-dependent translocation of PLC-ε to the plasma membrane. GTP loading of Rap2B by EGF was inhibited by chelation of intracellular Ca2+ and expression of lipase-inactive PLC-γ1 but not of PLC-ε. Expression of RasGRP3, a Ca2+/diacylglycerol-regulated guanine nucleotide exchange factor for Ras-like GTPases, but not expression of various other exchange factors enhanced GTP loading of Rap2B and PLC/Ca2+ signaling by the EGF receptor. EGF induced tyrosine phosphorylation of RasGRP3, but not RasGRP1, apparently caused by c-Src; inhibition of c-Src interfered with EGF-induced Rap2B activation and PLC stimulation. Collectively, these data suggest that the EGF receptor triggers activation of Rap2B via PLC-γ1 activation and tyrosine phosphorylation of RasGRP3 by c-Src, finally resulting in stimulation of PLC-ε.

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