Epidermal growth factor induces phosphorylation of extracellular signal-regulated kinase 2 via multiple pathways.

Abstract

Expression of p21'n-'7, a dominant negative mutant of p2l' that blocks p2l' activation by growth factors, inhibits activation of extracellular signal-regulated kinase 2 (ERK2) by insulin and platelet-derived growth factor in rat-1 cells [A. M. M. de Vries-Smits, B. M. T. Burgering, S. J. Leevers, C. J. Marshall, and J. L. Bos, Nature (London) 357:602-604, 1992]. Here we report that expression of p21'asAsn-17 does not abolish epidermal growth factor (EGF)-induced phosphorylation of ERK2 in fibroblasts. Since EGF activates p2l' in these cells, this indicates that EGF induces a p21'-independent pathway for the phosphorylation of ERK2 as well. We investigated whether activation of protein kinase C (PKC) or increase in intracellular calcium could be involved in p21a-independent signaling. In rat-i cells, inhibition of either PKC, by prolonged 12-0-tetradecanoylphorbol-13-acetate (TPA) pretreatment, or calcium influx, by ethylene glycol-bis(1-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) pretreatment, did not abolish EGF-induced ERK2 phosphorylation. However, a combined inhibition of both p2l' and calcium influx, but not PKC, resulted in a complete inhibition of EGF-induced ERK2 phosphorylation. In contrast, in Swiss 3T3 cells, inhibition of both p2l' activation and TPA-sensitive PKC, but not calcium influx, inhibited EGF-induced ERK2 phosphorylation. These results demonstrate that in fibroblasts, EGF induces alternative pathways of ERK2 phosphorylation in a cell-type-specific manner.Activation of transmembrane signaling receptors by first messengers generates the formation of second messengers. These second messengers are implicated in the induction of a myriad of intracellular signals of various biochemical constitutions. A common factor of signaling by many mitogenic stimuli is the activation of microtubule-associated protein 2 (MAP2) kinase (for recent reviews, see references 10, 40, and 42). Two forms of MAP2 kinase have been purified from fibroblasts with apparent molecular weights of 42 and 44 kDa. The p42m''K and p44mAPK have been cloned and are also named, respectively, extracellular signal-regulated kinase 2 (ERK2) and 1 (ERK1), to reflect the diversity of signals that can regulate them (3). Activation of both ERK1 and ERK2 requires phosphorylation at neighboring threonine and tyrosine residues and dephosphorylation of activated ERK2 by serine/threonine phosphatases or tyrosine phosphatases results in inactive ERK2 (1, 16). For ERK2, the sites of phosphorylation are located within one tryptic peptide and have been assigned to .Since full activation of ERK2 requires both threonine and tyrosine phosphorylation, it has been suggested that integration of two signaling pathways is required for the activation of the kinase (1). However, it is now clear that ERK2 is part of a kinase/phosphatase cascade in which at least several serine/threonine kinases act upstream of ERK2. The MAP kinase kinase (MAPKK or MEK) that activates ERK2 and phosphorylates the threonine and tyrosine site of ERK2 has recently been clo...

EGF receptor binding and transformation by v-cbl is ablated by the introduction of a loss-of-function mutation from the Caenorhabditis elegans sli-1 gene

“…The p42/p44 MAP kinase proteins are also known to be tyrosine phosphorylated following EGF stimulation (Burgering et al, 1993;Li et al, 1994). However immunoblotting total cell lysates from EGF-stimulated cells with anti-crkII, anti-crkL, or anti-MAP kinase antibodies showed that while these proteins are well expressed in ®broblasts, they do not migrate with the same mobility as the 42 kD phosphoprotein seen in v-cbl immunoprecipitates (data not shown).…”

Kinases: positive and negative regulators of apoptosis

“…44,45,[62][63][64][65][66] Ras is capable of activating both Raf [67][68][69] and Mekk; 70 however, activation of the Erk pathway can also occur in the absence of Ras activation. 71 The pathways leading to Erk activation are reported to be triggered by receptor tyrosine kinases, 72,73 G-protein linked receptors, [74][75][76] PKC, 71,77,78 and calcium. 46,[79][80][81][82] Several lines of evidence suggest that the Erk pathway has an important role in modulating the survival of hematopoietic cells.…”

Superagonistic activation of ErbB-1 by EGF-related growth factors with enhanced association and dissociation rate constants

“…The mitogen-activated protein kinase (MAPK) family members p42 and p44 are rapidly phosphorylated upon activation of ErbB receptors by both Ras and protein kinase Cmediated pathways (24). Previous work from our group has shown a direct correlation between MAPK activation and mitogenic activity of EGF-like growth factors (15).…”