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PubMed2000

Histone binding protein RbAp48 interacts with a complex of CREB binding protein and phosphorylated CREB.

A CREB-CREB binding protein (CBP) complex was used as bait to screen a mouse embryo cDNA library in yeast. One of the strongest interactions identified the histone binding protein RbAp48. RbAp48 also interacted weakly with CBP alone but did not interact with phosphorylated or nonphosphorylated CREB. CBP (or its homologue p300) from HeLa cell nuclear extracts coimmunoprecipitated with RbAp48 and its homologue RbAp46 and bound to a glutathione S-transferase-RbAp48 fusion protein. This interaction was stimulated by the addition of phosphorylated CREB and allowed the association of core histones and mononucleosomes in an acetylation-dependent manner. RbAp48 lowered the K(m) of CBP histone acetylase activity and facilitated p300-mediated in vitro transcription of a chromatinized template in the presence of acetylcoenzyme A. These data indicate that the association of phosphorylated CREB with CBP promotes the binding of RbAp48 and its homologue RbAp46, allowing the formation of a complex that facilitates histone acetylation during transcriptional activation.

Acetyl Coenzyme A↗
PubMed2001

Recruitment of an RNA polymerase II complex is mediated by the constitutive activation domain in CREB, independently of CREB phosphorylation.

The cAMP response element binding protein (CREB) is a bifunctional transcription activator, exerting its effects through a constitutive activation domain (CAD) and a distinct kinase inducible domain (KID), which requires phosphorylation of Ser-133 for activity. Both CAD and phospho-KID have been proposed to recruit polymerase complexes, but this has not been directly tested. Here, we show that the entire CREB activation domain or the CAD enhanced recruitment of a complex containing TFIID, TFIIB, and RNA polymerase II to a linked promoter. The nuclear extracts used mediated protein kinase A (PKA)-inducible transcription, but phosphorylation of CRG (both of the CREB activation domains fused to the Gal4 DNA binding domain) or KID-G4 did not mediate recruitment of a complex, and mutation of the PKA site in CRG abolished transcription induction by PKA but had no effect upon recruitment. The CREB-binding protein (CBP) was not detected in the recruited complex. Our results support a model for transcription activation in which the interaction between the CREB CAD and hTAFII130 of TFIID promotes the recruitment of a polymerase complex to the promoter.

Animals↗
PubMed1993

Structural determinants outside of the leucine zipper influence the interactions of CREB and ATF-2: interaction of CREB with ATF-2 blocks E1a-ATF-2 complex formation.

Dimerization of leucine zipper-containing proteins has been associated characteristically with the formation of a coiled-coil structure between two compatible leucine zipper motifs. In the present study we demonstrate the association of the leucine zipper of cAMP response element-binding protein (CREB) with a zinc finger motif of ATF-2. The association of the CREB leucine zipper with the ATF-2 zinc finger is stabilized if the ATF-2 leucine zipper is intact, implying that the preferred interactive structure of ATF-2 juxtaposes the amino-terminal zinc finger motif of this protein with the carboxy-terminal leucine zipper of this same protein. Furthermore, we demonstrate that the association of the CREB leucine zipper with the ATF-2 zinc finger in vitro blocks the association of the adenoviral E1a protein with ATF-2. Similarly, overexpression of full-length CREB, or a truncated version of this protein corresponding to the carboxy-terminal 74 amino acids that make up the DNA-binding and dimerization domains, can block the ATF-2-mediated transcriptional stimulation by E1a in vivo. Mutation of the ATF-2 zinc finger motif stimulates DNA binding of this protein, and abolishes interactions with E1a and CREB proteins. These results demonstrate that the structural conformation of ATF-2 is critical for DNA binding and protein-protein interactions and, further, that leucine zippers can mediate protein-protein interactions with structural motifs other than leucine zippers.

Activating Transcription Factors↗
PubMed2001

Cyclic AMP response element binding protein (CREB) and CREB binding protein (CBP) in global cerebral ischemia.

Cyclic AMP (cAMP) response element binding protein (CREB) is a transcription factor that has been implicated in neuronal responses to ischemia. We examined the effect of global cerebral ischemia in the rat on the expression of CREB, its transcriptionally active phosphorylated form (pCREB), and the nuclear adaptor protein, CREB binding protein (CBP). Global ischemia induced the expression of pCREB and CBP in vulnerable neurons of the hippocampal CA1 sector. In primary cultures of murine cortical neurons subjected to hypoxia, CBP was selectively expressed in cells with morphologically intact cell nuclei, and not in cells with condensed or fragmented nuclei indicative of irreversibly damaged neurons. These results support a role for transcriptional activation by CREB and CBP in neuronal cell-survival programs following cerebral ischemia.

Animals↗
PubMed2005

Activity-dependent neuroprotection and cAMP response element-binding protein (CREB): kinase coupling, stimulus intensity, and temporal regulation of CREB phosphorylation at serine 133.

The dual nature of the NMDA receptor as a mediator of excitotoxic cell death and activity-dependent cell survival likely results from divergent patterns of kinase activation, transcription factor activation, and gene expression. To begin to address this divergence, we examined cellular and molecular signaling events that couple excitotoxic and nontoxic levels of NMDA receptor stimulation to activation of the cAMP response element-binding protein (CREB)/cAMP response element (CRE) pathway in cultured cortical neurons. Pulses (10 min) of NMDA receptor-mediated synaptic activity (nontoxic) triggered sustained (up to 3 h) CREB phosphorylation (pCREB) at serine 133. In contrast, brief stimulation with an excitotoxic concentration of NMDA (50 microm) triggered transient pCREB. The duration of pCREB was dependent on calcineurin activity. Excitotoxic levels of NMDA stimulated calcineurin activity, whereas synaptic activity did not. Calcineurin inhibition reduced NMDA toxicity and converted the transient increase in pCREB into a sustained increase. In accordance with these observations, sustained pCREB (up to 3 h) did not require persistent kinase pathway activity. The sequence of stimulation with excitotoxic levels of NMDA and neuroprotective synaptic activity determined which stimulus exerted control over pCREB duration. Constitutively active and dominant-negative CREB constructs were used to implicate CREB in synaptic activity-dependent neuroprotection against NMDA-induced excitotoxicity. Together these data provide a framework to begin to understand how the neuroprotective and excitotoxic effects of NMDA receptor activity function in an antagonistic manner at the level of the CREB/CRE transcriptional pathway.

Action Potentials↗
PubMed1996

[Transcriptional regulation by CREB and proteins of CREB family].

CREB (cAMP-responsive element binding protein), which can be activated after phosphorylation by protein kinase A, plays an important role in cAMP-induced gene expression. Several recent studies have suggested that a co-activator designated as CREB binding protein (CBP) is crucial in mediating the transcriptional activity of CREB. In nervous system, in addition to playing a role in neurotransmitter-induced gene transcription, CREB may take part in mediating neurotrophin signals that ultimately lead to such cellular responses as proliferation, differentiation and survival.

Activating Transcription Factor 1↗
PubMed2002

Roles of phosphorylation and helix propensity in the binding of the KIX domain of CREB-binding protein by constitutive (c-Myb) and inducible (CREB) activators.

cAMP-response element-binding protein (CREB)-binding protein (CBP) is a general transcriptional co-activator that mediates interactions between transcription factors and the basal transcription machinery. To obtain insights into the mechanism by which the KIX domain of CBP can recognize the transactivation domains of many different transcription factors, we have used NMR and biochemical analyses to study the interactions of KIX with the transactivation domain from the constitutive activator c-Myb and with the kinase-inducible transactivation domain (KID) from CREB. NMR chemical shift mapping shows that both activation domains bind to the same surface of KIX. In the unbound state, both the phosphorylated KID and c-Myb activation domains are only partly structured, and binding to KIX is coupled with folding to form an amphipathic helix. Helix-destabilizing mutations significantly impair binding, whereas mutations that increase the intrinsic secondary structure content of the free phosphorylated KID peptide have only a small influence on binding affinity. Low affinity but specific binding of unphosphorylated KID to KIX was measured by ITC and was also observed in Western blot assays and by a fluorescence resonance energy transfer experiment in living cells. The large increase in the affinity for phosphorylated KID is due to favorable intermolecular interactions involving the phosphate moiety. After induction by phosphorylation, CREB is able to compete effectively with other transcriptional activators for binding to CBP.

Amino Acid Sequence↗
PubMed2005

Differential activation of the transcription factor cyclic AMP response element binding protein (CREB) in macrophages following infection with pathogenic and nonpathogenic mycobacteria and role for CREB in tumor necrosis factor alpha production.

Previous studies in our laboratory have shown a differential activation of the mitogen-activated protein kinases (MAPKs) in primary bone marrow-derived macrophages following infection with pathogenic Mycobacterium avium compared to the activation following infection with nonpathogenic Mycobacterium smegmatis. Additionally, M. smegmatis-infected macrophages produced significantly elevated levels of tumor necrosis factor alpha (TNF-alpha) compared to the levels produced by M. avium-infected macrophages. The TNF-alpha production was dependent on both p38 and extracellular signal-regulated kinase 1/2 (ERK 1/2) activation. However, the macrophage transcription factors downstream of the MAPKs, which were required for TNF-alpha production, remained undefined. In this study we determined that the transcription factor cyclic AMP response element binding protein (CREB) is significantly more activated in M. smegmatis-infected macrophages than in M. avium-infected macrophages. We also found that CREB activation was dependent on p38 and protein kinase A but not on ERK 1/2 or calmodulin kinase II. Moreover, mutating the cAMP-responsive element on the TNF-alpha promoter resulted in significantly diminished promoter activity following M. smegmatis infection but not M. avium infection. The inability of macrophages infected with M. avium to sustain MAPK activation and to produce high levels of TNF-alpha was due, in part, to an increase in serine/threonine phosphatase PP2A activity. Our studies are the first to demonstrate an important role for the transcription factor CREB in TNF-alpha production by mycobacterium-infected macrophages, as well as a role for M. avium's induction of PP2A phosphatase activity as a mechanism to limit macrophage activation.

Animals↗
PubMed2005

Coordinate regulation of forskolin-induced cellular proliferation in macrophages by protein kinase A/cAMP-response element-binding protein (CREB) and Epac1-Rap1 signaling: effects of silencing CREB gene expression on Akt activation.

In this study, we have examined the role of two cAMP downstream effectors protein kinase A (PKA) and Epac, in forskolin-induced macrophage proliferation. Treatment of macrophages with forskolin enhanced [(3)H]thymidine uptake and increased cell number, and both were profoundly reduced by prior treatment of cells with H-89, a specific PKA inhibitor. Incubation of macrophages with forskolin triggered the activation of Akt, predominantly by phosphorylation of Ser-473, as measured by Western blotting and assay of its kinase activity. Akt activation was significantly inhibited by LY294002 and wortmannin, specific inhibitors of phosphatidylinositol 3-kinase, but not by H-89. Incubation of macrophages with forskolin also increased Epac1 and Rap1.GTP. Immunoprecipitation of Epac1 in forskolin-stimulated cells co-immunoprecipitated Rap1, p-Akt(Thr-308), and p-Akt(Ser-473). Silencing of CREB gene expression by RNA interference prior to forskolin treatment not only decreased CREB protein and its phosphorylation at Ser-133, but also phosphorylation of Akt at Ser-473, and Thr-308. Concomitantly, this treatment inhibited [(3)H]thymidine uptake and reduced forskolin-induced proliferation of macrophages. Forskolin treatment also inhibited activation of the apoptotic mechanism while promoting up-regulation of the anti-apoptotic pathway. We conclude that forskolin mediates cellular proliferation via cAMP-dependent activation of both PKA and Epac.

Androstadienes↗
PubMed1992

Beta-adrenergic stimulation of cFOS via protein kinase A is mediated by cAMP regulatory element binding protein (CREB)-dependent and tissue-specific CREB-independent mechanisms in corticotrope cells.

Catecholamines stimulate proopiomelanocortin (POMC) gene expression in corticotrope cells, but the molecular mechanisms of these effects are not known. While beta-adrenergic receptors stimulate the protein kinase A (PKA) system, the POMC promoter does not have classical cAMP-response elements (CREs). Therefore, we investigated the induction of the c-fos protooncogen, previously shown to increase POMC transcription in AtT20 cells. In this corticotrope-derived cell line, we show that activation of beta-receptors with isoprenaline (Iso) induces a transient rise in c-fos mRNA levels. Gel mobility shift assays with a labeled AP1 consensus sequence (TGACTCA) showed induction of specific binding activity after Iso treatment. Cotransfection experiments with dominant inhibitory PKA mutants and reporter genes containing c-fos promoter sequences showed that c-fos induction by Iso is entirely dependent on a functional PKA activity. Furthermore, we show that beta-receptor induction of c-fos in corticotrophs is mediated by at least two distinct cAMP-responsive sequences. cAMP regulatory element binding (CREB)-dependent induction is observed on the CRE located at -60 bp on the c-fos promoter. A region located in the vicinity of the dyad symetry element (-290) is also found to mediate tissue-specific cAMP induction. Transcriptional activation by this site, although sensitive to PKA antagonism, is not blocked by CREB mutants.

Animals↗
PubMed1997

Somatostatin acts by inhibiting the cyclic 3',5'-adenosine monophosphate (cAMP)/protein kinase A pathway, cAMP response element-binding protein (CREB) phosphorylation, and CREB transcription potency.

Somatostatin (SRIF) was discovered as an inhibitor of GH secretion from pituitary somatotroph cells. SRIF analogs are very effective agents used to treat neuroendocrine tumors and are now being used with increasing frequency in clinical trials to treat more aggressive malignancies. However, the cellular components mediating SRIF signal transduction remain largely unknown. We have stably overexpressed the SRIF type 2 receptor (SST2) in GH4 rat somatomammotroph cells, establishing a physiologically relevant model system. In this model, the SRIF analog, BIM23014, inhibited forskolin-induced cAMP accumulation, protein kinase A activation, cAMP response element-binding protein phosphorylation, and Pit-1/GHF-1 promoter activation in an okadaic acid-insensitive manner. Pertussis toxin inhibited the effects of BIM23014, documenting that SST2 signaling was coupled to Gi. Moreover, the inhibitory effects of BIM23014 were reversed by overexpression of protein kinase A catalytic subunit, indicating that SRIF does not act via serine/threonine phosphatases, but, rather, by lowering protein kinase A activity. These data define the components of the SRIF/SST2 receptor signaling pathway and provide important mechanistic insights into how SRIF controls neuroendocrine tumors. As SRIF analogs are effective antitumor agents, and many other related compounds are in development, the knowledge gained here will further our understanding of their mechanism of action in other malignancies as well.

Animals↗
PubMed2006

Phosphorylation-dependent degradation of transgenic CREB protein initiated by heterodimerization.

The transcription factor CREB (cyclic AMP response element binding protein) is implicated in diverse brain functions and represents a prospective target in gene therapy for human disorders. However, the transgenic expression and stability of exogenously expressed CREB within the cell remains poorly characterized. Here we found that transient expression of a CREB dominant interfering mutant A-CREB or the inducible cAMP early repressor, ICER, led to the dramatic decrease of exogenously co-expressed CREB in 293 human embryonic kidney cells. Elevation of protein kinase A activity within the cells restored CREB protein levels. A-CREB did not effect the transient expression of a truncated CREB lacking the leucine zipper domain demonstrating a specific effect of heterodimerization on CREB protein stability. Somatic gene transfer into the rat brain using a recombinant adeno-associated virus vector provided robust expression of both transgenic CREB and ICER mRNAs under the control of a constitutive neuron specific enolase (NSE) promoter. In contrast to ICER, the expression of the transgenic CREB mRNA did not result in elevation of CREB protein levels within dentate granule cells of the hippocampus, suggesting its prompt degradation under basal conditions. However, following tetanization of the perforant pathway, which is known to induce CREB phosphorylation, there was a significant increase in the amount of transgenic CREB protein within dentate granule cells. Hence, heterodimerization of unphosphorylated CREB with either A-CREB or ICER triggers CREB protein degradation, whereas phosphorylation prevents CREB from such degradation both in vitro and in vivo.

Animals↗
PubMed2004

Colocalization of phosphorylated CREB with calcium/calmodulin-dependent protein kinase IV in hippocampal neurons induced by ohmfentanyl stereoisomers.

The transcription factor cAMP response element-binding protein (CREB) plays an important role in opioids dependence. To better understand the role of CREB in opioids dependence and underlying signal pathways, we compared the effects of three ohmfentanyl stereoisomers ((-)-cis-(3R,4S,2'R) OMF (F9202), (+)-cis-(3R,4S,2'S) OMF (F9204), (-)-cis-(3S,4S,2'R) OMF (F9203)) and morphine on CREB phosphorylation and the expression of Ca2+/calmodulin-dependent protein kinase IV (CaMKIV) in hippocampus derived from mice which displayed conditioned place preference (CPP) behavior by Western blot, and immunohistochemistry analyses. Moreover, we studied the effects of OMF and morphine on CREB phosphorylation and colocalization of phosphorylated CREB (P-CREB) with CaMKIV in cultured rat hippocampal neurons by Western blot, and confocal fluorescence microscopy analyses. The results showed that F9202, F9204 or morphine, which could induce CPP, enhanced CREB phosphorylation and the expression of CaMKIV in hippocampus from CPP mice without affecting total CREB protein level. The CREB phosphorylation of cultured hippocampal neurons was also enhanced and reached its peak level at 30 min upon exposure to F9202 (100 nM), F9204 (100 nM) or morphine (1 microM), while the total CREB protein level was not altered. KN-62 (10 microM), an inhibitor of CaM kinases, prevented CREB phosphorylation induced by morphine, F9202, and F9204 without change of total CREB level. The results of confocal fluorescence microscopy further demonstrated that the activated CREB (P-CREB) was colocalized with CaMKIV in nucleus. F9203, which could not induce CPP, failed to increase the CREB phosphorylation and the colocalization of P-CREB with CaMKIV both in hippocampus from CPP mice and in cultured hippocampal neurons. This is the first evidence to suggest that the increased CREB phosphorylation via CaMKIV signal pathway in hippocampus is relevant to opioids psychological dependence.

Animals↗
PubMed1995

cAMP response element-binding protein (CREB) interacts with transcription factors IIB and IID.

cAMP response element-binding protein (CREB) participates in both constitutive and cAMP-induced transcription of cAMP-responsive genes. CREB-mediated constitutive transcription requires only CREB-binding sites and a minimal promoter region (containing the TATA through start sequences), indicating that CREB interacts directly with components of the general transcription machinery. In this study, a coimmunoprecipitation assay was used to test for interaction of CREB with the general transcription factors (TF) TFIIB and TFIID and the core component of TFIID, TATA-binding protein (TBP). Human TFIIB and TBP, tagged with distinct epitopes (eTFIIB and eTBP), were expressed in and purified from Escherichia coli, and holo-eTFIID, containing eTBP, was obtained from the HeLa cell line LTR alpha 3. 35S-Labeled CREB, synthesized in vitro and incubated with eTFIIB, was coimmunoprecipitated with antibody recognizing eTFIIB, indicating that CREB specifically binds to TFIIB. 35S-CREB was coimmunoprecipitated with antibody against eTBP, but only when incubated with the holo-eTFIID complex, not with eTBP alone. TFIIB interacted with TBP, but CREB was not coprecipitated with the eTBP antibody when incubated with eTBP plus TFIIB, so CREB did not form a stable ternary complex with TFIIB and TBP. Conversely, depletion of TFIIB from the holo-TFIID preparation did not diminish the level of interaction between CREB and TFIID. Thus, CREB interacts independently with TFIIB and TFIID, but not directly with TBP. A protein kinase A phosphorylation site mutant of CREB and wild-type CREB exhibited equivalent interaction with TFIIB, indicating that this phosphorylation is not required. Consistent with the role of CREB in promoting constitutive or basal transcription, the constitutive activation domain of CREB was sufficient for interaction with both TFIIB and TFIID.

Cyclic AMP Response Element-Binding Protein↗
PubMed1994

Melatonin regulates the phosphorylation of CREB in ovine pars tuberalis.

This study investigated whether melatonin could modulate the phosphorylation of the calcium/cyclic AMP response-element binding-protein (CREB) within primary cell cultures of ovine pars tuberalis (oPT) and pars distalis (oPD). Gel shift assays confirmed the presence of nuclear factors able to alter the electrophoretic mobility of a 32P-labelled CRE oligonucleotide. Two shifted bands were observed probably due to monomer and dimer binding to the CRE. Each band was supershifted by antisera directed against both CREB and the phosphorylated form of CREB (P-CREB), consistent with a specific role of CREB proteins in transcriptional regulation. To study the physiological role of CREB, the nuclear immunoreactivity for P-CREB was followed in primary cultures of oPT given different pharmacological treatments. Cells stimulated with forskolin responded with a robust time- and dose-dependent increase in nuclear phospho-CREB immunoreactivity (P-CREB-ir), confirming that activation of this transcription factor occurred through the cyclic AMP-PKA pathway. Maximal stimulation was achieved within 15 min and persisted for up to 1 h. Treatment with melatonin alone did not alter basal P-CREB-ir levels, yet melatonin inhibited the forskolin-induced increase in P-CREB-ir in a dose-dependent manner (IC50 of between 10(-10) M and 10(-8) M melatonin when tested against 1 microM forskolin). In contrast, in primary cultures of oPD, melatonin failed to block forskolin-stimulated increases in either the content of cyclic AMP or the intensity of nuclear P-CREB-ir, confirming that the action of melatonin upon P-CREB-ir is tissue specific. These results demonstrate that, consistent with its inhibitory effect on the activation of PKA within oPT, melatonin prevents or reverses the phosphorylation of CREB induced by activation of the cyclic AMP signal transduction pathway. Therefore melatonin has the potential to regulate gene expression in the oPT by acting upon the CREB transcription factor. However, this paper also shows that 12-O-tetradecanoylphorbol-13-acetate (TPA) which activates PKC also leads to the phosphorylation of CREB in oPT cells, suggesting the potential involvement of other signal transduction pathways in the transcriptional regulation of these cells.

Animals↗
PubMed2005

CREB mediates ERK-induced survival of mouse renal tubular cells after oxidant stress.

BACKGROUND: We showed that extracellular signal-regulated protein kinase (ERK) is prosurvival during oxidant stress both in the kidney and in cultured mouse proximal tubule (TKPTS) cells and demonstrated concomitant activation of ERK as well as the cyclic adenosine monophosphate (cAMP)-responsive element binding protein (CREB), during survival in vitro. We now show that CREB is a necessary prosurvival target of ERK. METHODS: Ischemia/reperfusion (I/R) injury was induced in 129Sv mice. Oxidant stress was induced by hydrogen peroxide (H(2)O(2)) in TKPTS cells. Activation of CREB was determined by immunohistochemistry and Western blotting. Inhibition and activation of CREB was achieved by mutant or activated CREB-containing adenoviruses in vitro. The effects of oxidant stress on cell survival, CREB binding, and CREB-mediated transcription was determined by cell counting, gelshift analysis, and luciferase assay, respectively. RESULTS: I/R activates CREB in the surviving distal nephron segments of the kidney. Inhibition of ERK and CREB abrogates survival after 0.5 mmol/L H(2)O(2) treatment, while overexpression of CREB ameliorates necrotic death caused by 1 mmol/L H(2)O(2). Inhibition of ERK also inhibited CREB activation. Binding of phosphorylated CREB to a CREB oligonucleotide was significantly increased after 0.5 mmol/L H(2)O(2) but decreased after 1 mmol/L H(2)O(2). Similarly, CREB-mediated transcription was significantly increased after 0.5 mmol/L H(2)O(2) treatment, while 1 mmol/L H(2)O(2) inhibited it. Interestingly, transcription from the CREB-driven bcl-2 promoter was unchanged after 0.5 mmol/L but decreased after 1 mmol/L H(2)O(2) treatment in agreement with Western blot studies. CONCLUSION: We show that survival during oxidant stress is mediated through CREB and identification of its downstream targets will reveal important survival pathways.

Animals↗
PubMed1994

A monomeric derivative of the cellular transcription factor CREB functions as a constitutive activator.

The mammalian transcriptional activator CREB binds as a dimer to a broad spectrum of inducible promoters. CREB activity is modulated by several signalling agents (protein kinase A [PKA], Ca2+, and transforming growth factor beta) and via functional interactions with cell-specific transcription factors. In addition, CREB can activate transcription constitutively and repress the activity of several other transcriptional activators. The mechanisms that allow CREB to act in such a malleable manner and the role that CREB dimerization might play in this are poorly understood. To probe the latter issue, we have created monomeric forms of CREB by fusing CREB to the DNA-binding domain of a protein (B-cell specific activator protein [BSAP]) that binds to DNA as a monomer. Remarkably, monomeric CREB acts as a potent, constitutive activator under conditions in which native CREB is inducible by PKA. Thus, CREB contains constitutive activation regions that are unable to function in native CREB. Two glutamine-rich domains that are important for native, PKA-inducible CREB activity are required for the constitutive activity of monomeric CREB. In contrast, two elements within the kinase-inducible domain of CREB are dispensable for constitutive activity. We discuss our results in relation to inducible and constitutive CREB activity and the potential modes of action of other activators that directly interact with CREB.

Base Sequence↗
PubMed2000

Blockade of cyclic AMP-responsive element DNA binding in the brain of CREB delta/alpha mutant mice.

The cAMP-responsive element binding protein (CREB) gene transcription factor has been implicated in the synaptic plasticity and memory. Here, we investigated the mechanisms of CREB and/or cyclic AMP-responsive element modulatory protein (CREM) binding to CRE sites in brain tissues. CRE-DNA binding was determined in nuclear extracts obtained from the several brain structures of wild-type and CREB delta/alpha mutant mice. It was found that antibodies to CREB, phosphorylated CREB, and CREM supershifted the CRE-DNA binding complex in cortical nuclear extracts from wild-type mice, which suggests that the CRE-DNA binding complex contains both CREB and CREM proteins. In contrast, CRE-DNA binding is abolished in the cortex, hippocampus, cerebellum, and amygdala of CREB delta/alpha mutant mice. Because the CREB delta and alpha isoforms have been deleted in CREB mutant mice, consequently, other forms of CREB, such as CREB-beta and CREM, are up-regulated. These results suggest that the binding of CREM to CRE sites requires the presence of CREB delta/alpha, and that CREB-beta may be inefficient in binding to CRE-sites. Thus, these results indicate that CREB delta/alpha mutant mice are a useful animal model for studying the functional role of CREB-dependent gene expression.

Animals↗