Stimulation of c-Rel transcriptional activity by PKA catalytic subunit ?

Abstract

Nuclear factor kappaB (NF-kappaB) is a eukaryotic transcription factor which responds to different extracellular signals. It is involved in immune response, inflammation, and cell proliferation. Increased expression of c-Rel (or its viral homolog v-Rel), one component of the NF-kappaB factors, induces tumorigenesis in different systems. The activity of NF-kappaB can be regulated by protein kinase A (PKA) in a cAMP-independent manner. Our previous results showed that c-MYC induces the activity of PKA by inducing the transcription of the gene encoding the PKA catalytic subunit beta (PKA-Cbeta). Constitutive expression of PKA-Cbeta in Rat1a cells induces their transformation. Here we show that CREB is unlikely to be a phosphorylation target of PKA-Cbeta as characterized by different cell lines. Electrophoretic mobility shift assays showed that c-Rel is present as a significant component of the NF-kappaB factors in c-MYC overexpressing status. The transcriptional activity of c-Rel was significantly stimulated by PKA-Cbeta. Coactivators p300/CBP are at least partially responsible for the enhanced activation mediated by c-Rel and PKA-Cbeta. Interaction between c-Rel and PKA-Cbeta was demonstrated using coimmunoprecipitation assays. Immunoprecipitation-in vitro phosphorylation assays showed the direct phosphorylation of c-Rel by PKA-Cbeta. These results indicate that c-Rel is a reasonable phosphorylation target of PKA-Cbeta, and that the transcriptional activity of c-Rel is stimulated by PKA-Cbeta possibly through the interaction with p300/CBP.

Thrombin and Collagen Induce a Feedback Inhibitory Signaling Pathway in Platelets Involving Dissociation of the Catalytic Subunit of Protein Kinase A from an NFκB-IκB Complex

“…Thrombin-induced NFB activation in both platelets and HL-60 cells (used as a positive control) was demonstrated by IB␣ phosphorylation and degradation, and by increased DNA-binding of activated NFB. We could show that in platelets, like in other cell types (15,17,47), a portion of PKAc binds to IB␣ and co-immunoprecipitates with IB␣. This interaction was significantly reduced in thrombin-and collagen-stimulated platelets, indicating release of catalytically active PKAc from IB␣ (Fig.…”

“…Elevation of cAMP levels and binding of cAMP to PKAr causes dissociation of the kinase complex and release of free active catalytic subunits (11)(12)(13)(14). However, in addition to this "classical" cAMP-dependent regulation of PKA activity, cAMP-independent activation of PKA has been demonstrated in different cell types (15)(16)(17). Some portion of PKAc molecules (independently from PKAr) is bound to IB in an NFB-IB complex.…”

Post-translational modifications regulating the activity and function of the nuclear factor kappa B pathway

“…p50 and c-Rel are also modified by PKAc at Ser-337 and Ser-267, respectively, the equivalent sites to RelA Ser-276 (Mosialos et al, 1991;Hou et al, 2003;Yu et al, 2004;Guan et al, 2005) (Figure 3; Table 3). Since this serine is also conserved in RelB (but not p52), it is probable that similar modification occurs with this subunit.…”

“…Ser-276 in RelA is also phosphorylated by the MSK1 kinase, although this is a nuclear event (see below) (Vermeulen et al, 2003). Phosphorylation at Ser-276 (and the analogous site in c-Rel) is thought to disrupt an intramolecular interaction between the RelA N and C termini, allowing DNA binding and p300/CBP coactivator interaction (Mosialos et al, 1991;Zhong et al, 1998;Zhong et al, 2002;Yu et al, 2004). However, the analysis of relaÀ/À MEFs reconstituted with S276A mutant versions of RelA indicates that the effect of phosphorylation of this site can be very promoter specific (Anrather et al, 2005), suggesting other effects might compensate for lack of phosphorylation at this site (see below) or that Ser-276 phosphorylation has a subtler effect on RelA function in cells.…”

Mutation of an IKK phosphorylation site within the transactivation domain of REL in two patients with B-cell lymphoma enhances REL's in vitro transforming activity

“…The C-terminal half of REL (aa 296-587) contains a transactivation domain, which is comprised of at least two subdomains: subdomain I (aa 422-497) and subdomain II (aa 518-587) (Martin et al, 2001;Starczynowski et al, 2003). The REL C-terminal transactivation domain can also be regulated by phosphorylation of specific serine (Ser) residues (Fognani et al, 2000;Martin and Fresno, 2000;Martin et al, 2001;Yu et al, 2004;Lawrence et al, 2005;Harris et al, 2006). Recently, IkB kinase (IKK)-associated kinases have been found to phosphorylate C-terminal residues of REL in an immune complex kinase assay (Lawrence et al, 2005;Harris et al, 2006).…”