cAMP Activates MAP Kinase and Elk-1 through a B-Raf- and Rap1-Dependent Pathway

Abstract

Cyclic adenosine monophosphate (cAMP) has tissue-specific effects on growth, differentiation, and gene expression. We show here that cAMP can activate the transcription factor Elk-1 and induce neuronal differentiation of PC12 cells via its activation of the MAP kinase cascade. These cell type-specific actions of cAMP require the expression of the serine/threonine kinase B-Raf and activation of the small G protein Rap1. Rap1, activated by mutation or by the cAMP-dependent protein kinase PKA, is a selective activator of B-Raf and an inhibitor of Raf-1. Therefore, in B-Raf-expressing cells, the activation of Rap1 provides a mechanism for tissue-specific regulation of cell growth and differentiation via MAP kinase.

The mitogen-activated protein (MAP) kinase cascade can either stimulate or inhibit DNA synthesis in primary cultures of rat hepatocytes depending upon whether its activation is acute/phasic or chronic

“…In contrast to NGF treatment, epidermal growth factor (EGF) treatment of PC12 cells induced an acute phasic activation of p42 MAPkinase which returned to baseline within 30 min ( Figure 1B). These data are in agreement with data of several other laboratories [16,21,22,34,35]. Bailie et al reported that primary cultures of hepatocytes possess binding sites for NGF, and we investigated whether NGF had a similar capability to modulate the MAP kinase cascade in these cells [26].…”

Biochemical characterization of C3G: an exchange factor that discriminates between Rap1 and Rap2 and is not inhibited by Rap1A(S17N)

“…For inhibition of the Ras pathway much smaller amounts of Ras(S17N) are normally used. More likely however, the dominant negative e ect of (S17N)Rap1B observed by Vossler et al is caused by titration of another not yet identi®ed Rap-GEF, since C3G could not potentiate the cAMP-mediated activation of ERKs (Vossler et al, 1997). In another very recent paper, C3G and Rap1 were also found to be on separate pathways (Tanaka et al, 1997).…”

“…mGDP in the presence or absence of the indicated concentrations of C3G-CD after addition of 100-fold excess of unlabeled GDP. The assay was performed at 258C in 50 mM Tris, pH, 7.5, 5 mM MgCl 2 and 5 mM DTE submitting our manuscript, Vossler et al (1997) described a dominant negative e ect of (S17N)Rap1A on the cAMP-dependent activation of Elk-1. This seems to be in contrast to our data, but one explanation could be the very large amount of (S17N)Rap1B (10 mg of DNA) that was used to transfect PC12 cells as we do observe some inhibition using 10 mM (S17N)Rap1A in our in vitro assay.…”

Quantitative determination of Rap 1 activation in cyclic nucleotide-treated HL-60 leukemic cells: lack of Rap 1 activation in variant cells

“…Assessing total cellular Rap 1 by immunoblotting is helpful but again only semiquantitative (Posern et al, 1998). Rap 1 activation has been assessed by other methods but these require transfecting cells with a tagged-version of Rap and/or incubating cells with high amounts of 32 PO 4 (Altschuler et al, 1995;Vossler et al, 1997;Boussiotis et al, 1997). In this work we describe a quantitative method to assess Rap 1 activation which does not require manipulation of the cells and we have applied this method to cultured cells; the method can also be used to assess Rap 1 activation in tissue samples.…”