Immediate early genes: their involvement in physiological and pathological responses in the nervous system.
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Animals↗
Biomedical subjects
T Curran
Publications and source records attributed to T Curran.
The regulation of c-fos: too much is never enough.
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DNA-Binding Proteins↗
A fos-lac Z transgenic mouse that can be used for neuroanatomic mapping.
Cellular immediate-early genes are rapidly induced by a diverse range of agents and conditions. Since many cIE genes encode known or potential transcription factors, they are believed to couple extracellular stimuli to long-lasting alterations in cellular phenotype through the regulation of gene transcription. In addition, the localization of the products of cIE genes has been used as a method for determining the cellular sites of action of particular agents in the nervous system. However, the methods of analysis are tedious, and the results may be ambiguous because of cross-reaction of reagents with related proteins. To further the utility of this approach, a bacterial gene encoding beta-galactosidase (lac Z) has been fused, in frame, into the fourth exon of c-fos, and this fos-lac Z fusion gene has been introduced into the germ line of mice. We have analyzed the expression of beta-galactosidase (under the control of the c-fos promoter) in the developing and adult nervous systems of these transgenic mice. As far as can be determined, the constitutive and stimulated expression of the transgene accurately reflects the expression of cognate c-fos in both cultured cells and the intact animal. This study has also revealed novel sites of constitutive and induced expression of c-fos that were overlooked using conventional analysis. In particular, constitutive expression of c-fos is associated with cells that are entering terminal differentiation and are destined to die. In addition, induced expression of the transgene in adult brain mirrors the pattern of neurotoxicity elicited by kainic acid.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Escape from redox regulation enhances the transforming activity of Fos.
Fos and Jun form dimeric complexes that bind to DNA sequences containing activator protein 1 (AP-1) sites and regulate gene expression. The in vitro DNA-binding activity of these proteins is sensitive to reduction-oxidation (redox). Reduction of a single conserved cysteine residue, located in the DNA-binding domain, either by reducing agents or by a nuclear redox factor (Ref-1), is required for AP-1 DNA-binding activity. Replacing the critical cysteine with serine results in a protein that can bind to DNA in vitro even under oxidizing conditions. To determine whether redox control affects the function of Fos in vivo, we have constructed, and compared the properties of, retroviral vectors expressing either a truncated Fos protein (F118-211) or a truncated Fos protein in which the critical cysteine was replaced by serine (FC154S). In infected chicken embryo fibroblasts (CEFs), both vectors expressed similar levels of Fos protein, which formed heterodimers with Jun at equivalent efficiencies. However, extracts from cells expressing FC154S exhibited a threefold increase in AP-1 DNA-binding activity compared with cells expressing F118-211. Furthermore, this enhanced binding activity was resistant to treatment with the oxidizing agent diamide. Infection of CEFs by virus expressing FC154S resulted in increased numbers of transformed colonies and an increase in colony size compared with those obtained following infection by virus expressing Fos 118-211. These results suggest that redox regulation may limit the total level of functional Fos-Jun complexes in vivo and that escape from this control enhances transforming activity.
Animals↗
Identification and characterization of Ref-1, a nuclear protein that facilitates AP-1 DNA-binding activity.
Fos and Jun form a heterodimeric complex that regulates gene transcription by binding to the activator protein-1 (AP-1) DNA sequence motif. Previously, we demonstrated that the DNA-binding activity of Fos and Jun is regulated in vitro by a novel redox (reduction-oxidation) mechanism. Reduction of a conserved cysteine (cys) residue in the DNA-binding domains of Fos and Jun by chemical reducing agents or by a nuclear redox factor stimulates DNA-binding activity. Here, we describe purification and characterization of a 37 kDa protein (Ref-1) corresponding to the redox factor. Although Ref-1 does not bind to the AP-1 site in association with Fos and Jun, it partially copurifies with a subset of AP-1 proteins. Purified Ref-1 protein stimulates AP-1 DNA-binding activity through the conserved Cys residues in Fos and Jun, but it does not alter the DNA-binding specificity of Fos and Jun. Ref-1 may represent a novel redox component of the signal transduction processes that regulate eukaryotic gene expression.
Cell Nucleus↗
Redox activation of Fos-Jun DNA binding activity is mediated by a DNA repair enzyme.
The DNA binding activity of Fos and Jun is regulated in vitro by a post-translational mechanism involving reduction-oxidation. Redox regulation occurs through a conserved cysteine residue located in the DNA binding domain of Fos and Jun. Reduction of this residue by chemical reducing agents or by a ubiquitous nuclear redox factor (Ref-1) recently purified from Hela cells, stimulates AP-1 DNA binding activity in vitro, whereas oxidation or chemical modification of the cysteine has an inhibitory effect on DNA binding activity. Here we demonstrate that the protein product of the ref-1 gene stimulates the DNA binding activity of Fos-Jun heterodimers, Jun-Jun homodimers and Hela cell AP-1 proteins as well as that of several other transcription factors including NF-kappa B, Myb and members of the ATF/CREB family. Furthermore, immunodepletion analysis indicates that Ref-1 is the major AP-1 redox activity in Hela nuclear extracts. Interestingly, Ref-1 is a bifunctional protein; it also possesses an apurinic/apyrimidinic (AP) endonuclease DNA repair activity. However, the redox and DNA repair activities of Ref-1 can, in part, be distinguished biochemically. This study suggests a novel link between transcription factor regulation, oxidative signalling and DNA repair processes in higher eukaryotes.
Amino Acid Sequence↗
Temporal and spatial expression of a fos-lacZ transgene in the developing nervous system.
A Fos-lacZ transgenic mouse has been described that accurately recapitulates both constitutive and inducible patterns of c-fos expression in adult mice. Here we describe the developmental expression of the transgene in the brain during the early postnatal period. On the day of birth, expression of the transgene is observed in several discrete regions of the CNS; including the olfactory bulb, hippocampus, retrosplenial cortex, parafascicular nucleus of the thalamus, and several cranial nerve nuclei. In these regions, expression declines to adult levels by three weeks. In other regions of the CNS, expression appeared transiently after P0.
Animals↗
fos-lacZ transgenic mice: mapping sites of gene induction in the central nervous system.
A transgenic mouse line containing a fos-lacZ fusion gene was derived in which beta-galactosidase activity identified cell populations expressing fos either constitutively or after stimulation. Seizures and light pulses induced nuclear lacZ activity in defined populations of neurons in vivo, and an array of neurotransmitters, including glutamate, induced the transgene in primary brain cultures. In unstimulated mice, the major sites of fos-lacZ expression were skin, hair follicle, and bone. fos-lacZ mice provide a new avenue for activity mapping studies based on gene expression.
Adrenal Glands↗
Crossed signals: oncogenic transcription factors.
Transcription factors operate at key regulatory junctions in the cell's responses to diverse extracellular stimuli. We discuss how the transforming activity of mutated transcription factors encoded by several oncogenes (v-erbA, v-fos, v-jun and v-rel) may result in part from a loss of the integrated response to the signalling network.
Genes, fos↗
Effects of similarity and repetition on memory: registration without learning?
We investigated judgments of the frequency of test items (Y) that were highly similar to studied items (X) to test a prediction made by several memory models: that the judged frequency of Y should be proportional to the judged frequency of X. Whether stimuli were pictures or words, judged frequency of Y was bimodally distributed with 1 mode at zero, suggesting that frequency judgments involve a 2-stage process in which a zero judgment is made if there is a mismatch between retrieved information and the test item. Nonzero judgements, taken by themselves, were consistent with the prediction of proportionality. In 2 experiments, the percentage of zero judgments made to Y increased with repetition of X, but in 2 others the percentage did not change beyond frequency = 1. The percentage of "new" judgments in recognition memory followed this same pattern. Because the judged frequency of X increased even as X-Y discrimination showed no improvement, we characterize the result as "registration without learning."
Adult↗
Jun is phosphorylated by several protein kinases at the same sites that are modified in serum-stimulated fibroblasts.
c-jun is a member of the family of immediate-early genes whose expression is induced by factors such as serum stimulation, phorbol ester, and differentiation signals. Here we show that increased Jun synthesis after serum stimulation is accompanied by a concomitant increase in phosphorylation. Several serine-threonine kinases were evaluated for their ability to phosphorylate Jun in vitro. p34cdc2, protein kinase C, casein kinase II, and pp44mapk phosphorylated Jun efficiently, whereas cyclic AMP-dependent protein kinase and glycogen synthase kinase III did not. The sites phosphorylated by p34cdc2 were similar to those phosphorylated in vivo after serum induction. The major sites of phosphorylation were mapped to serines 63, 73, and 246. Phosphorylation of full-length Jun with several kinases did not affect the DNA-binding activity of Jun homodimers or Fos-Jun heterodimers. Comparison of the DNA binding and in vitro transcription properties of wild-type and mutated proteins containing either alanine or aspartic acid residues in place of Ser-63, -73, and -246 revealed only minor differences among homodimeric complexes and no differences among Fos-Jun heterodimers. Thus, phosphorylation of Jun did not produce a significant change in dimerization, DNA-binding, or in vitro transcription activity. The regulatory role of phosphorylation in the modulation of Jun function is likely to be considerably more complex than previously suggested.
Blood↗
Fos and Jun: oncogenic transcription factors.
The fos and jun proto-oncogenes are members of the set of genes known as cellular immediate-early genes. Their expression is induced transiently by a great variety of extracellular stimuli associated with mitogenesis, differentiation processes or depolarization of neurons. They encode DNA binding proteins that form dimeric complexes through a leucine zipper structure that function as transcription factors. Continuous overexpression of fos or jun causes transformation of fibroblasts. Because of their ubiquitous expression it is believed that the target genes regulated by Fos and Jun are different in the many circumstances in which they are expressed. Thus, their functional specificity is likely to be regulated at several levels. We have uncovered several potential mechanisms that could contribute to their regulation. These include formation of a large number of heterodimeric complexes, post-translational modification by phosphorylation and a novel reduction/oxidation (redox) mechanism, presence of both positive and negative transcriptional domains and the ability of Fos and Jun to induce distinct bends in DNA structure.
Animals↗
DNA bending by Fos and Jun: the flexible hinge model.
DNA bending is essential for the assembly of multiprotein complexes that contact several DNA sequence elements. An approach based on phasing analysis was developed that allows determination of both the directed DNA bend angle and the orientation of DNA bending. This technique has been applied to the analysis of DNA bending by the transcription regulatory proteins Fos and Jun. Complexes that contained different combinations of full-length and truncated Fos and Jun induced DNA bends of different magnitudes and orientations. The DNA bends induced by the individual proteins were determined on the basis of a quantitative model for DNA bending by dimeric complexes. This information was used to visualize the consequences of DNA bending by Fos and Jun for the structures of Fos-Jun-DNA and Jun-DNA complexes.
Computer Graphics↗
Pro-Leu-Ser/Thr-Pro is a consensus primary sequence for substrate protein phosphorylation. Characterization of the phosphorylation of c-myc and c-jun proteins by an epidermal growth factor receptor threonine 669 protein kinase.
A growth factor-stimulated (MAP2-related) protein kinase, ERT, that phosphorylates the epidermal growth factor receptor at Thr669 has been purified from KB human tumor cells by Northwood and co-workers (Northwood, I. C., Gonzalez, F. A., Wartmann, M., Raden, D. L., and Davis, R. J. (1991) J. Biol. Chem. 266, 15266-15276). The ERT protein kinase has a restricted substrate specificity, and the structural determinants employed for substrate recognition by this enzyme have not been defined. As an approach toward understanding the specificity of substrate phosphorylation, we have used an in vitro assay to identify additional substrates for the ERT protein kinase. In this report we describe two novel substrates: (a) the human c-myc protein at Ser62 and (b) the rat c-jun protein at Ser246. Alignment of the primary sequences surrounding the phosphorylation sites located within the epidermal growth factor receptor (Thr669), Myc (Ser62), and Jun (Ser246) demonstrated a marked similarity. The observed consensus sequence was Pro-Leu-Ser/Thr-Pro. We propose that this sequence forms part of a substrate structure that is recognized by the ERT protein kinase.
Amino Acid Sequence↗
Fos-Jun heterodimers and Jun homodimers bend DNA in opposite orientations: implications for transcription factor cooperativity.
Association of Fos and Jun with the AP-1 site results in a conformational change in the basic amino acid regions that constitute the DNA-binding domain. We show that Fos and Jun induce a corresponding alteration in the conformation of the DNA helix. Circular permutation analysis indicated that both Fos-Jun heterodimers and Jun homodimers induce flexure at the AP-1 site. Phasing analysis demonstrated that Fos-Jun heterodimers and Jun homodimers induce DNA bends that are directed in opposite orientations. Fos-Jun heterodimers bend DNA toward the major groove, whereas Jun homodimers bend DNA toward the minor groove. Fos and Jun peptides encompassing the dimerization and DNA-binding domains bend DNA in the same orientations as the full-length proteins. However, additional regions of both proteins influence the magnitude of the DNA bend angle. Thus, despite the amino acid sequence similarity in the basic region Fos-Jun heterodimers and Jun homodimers form topologically distinct DNA-protein complexes.
Base Sequence↗
Regulation of a fos-lacZ fusion gene: a paradigm for quantitative analysis of stimulus-transcription coupling.
Expression of the c-fos protooncogene is induced by a great variety of extracellular stimuli. A fos-lacZ fusion gene has been constructed that recapitulates this regulation. The fos-lacZ gene was introduced into B104 neuroblastoma cells for use in a quantitative assay for stimulus-transcription coupling. Both alpha- and beta-adrenergic agonists, dibutyryl cAMP, and phorbol ester induced beta-galactosidase activity in a dose-dependent manner. Thus, the interactions of receptors with agonists and antagonists, as well as intracellular second messenger-mediated signaling events, can be analyzed quantitatively. This approach represents a prototypic method for investigating stimulus-response coupling based upon gene expression.
Animals↗
Cross-family dimerization of transcription factors Fos/Jun and ATF/CREB alters DNA binding specificity.
The Fos/Jun and ATF/CREB families of transcription factors function in coupling extracellular signals to alterations in expression of specific target genes. Like many eukaryotic transcription factors, these proteins bind to DNA as dimers. Dimerization is mediated by a structure known as the "leucine-zipper" motif. Although Fos/Jun and ATF/CREB were previously thought to interact preferentially with different DNA regulatory elements (the AP-1/TRE and ATF/CRE sites, respectively), we find that members of these two families form selective cross-family heterodimers. The resulting heterodimers display distinguishable DNA binding specificities from each other and from their parental homodimers. These findings indicate that the Fos/Jun and ATF/CREB families of transcription factors are not as distinct as was previously thought. We suggest that they can be grouped into a superfamily of transcription factors.
Activating Transcription Factor 2↗
Proto-oncogene transcription factors and epilepsy.
Chemically and electrically induced seizures elicit the rapid transcriptional activation in neurons of a class of genes referred to as cellular immediate-early genes. Since the products of these genes include transcription factors and cytokines, they are proposed to be involved in coupling neuronal excitation to a complex, and poorly understood, programme of cellular responses that involves the regulation of gene expression. Products of two cellular immediate-early genes, c-fos and c-jun, are components of the transcription factor AP-1. In this review, Jim Morgan and Tom Curran discuss how these gene products have begun to reveal some of the molecular details of stimulus-transcription coupling in the nervous system following seizures. In addition, these genes have provided novel reagents and concepts for investigating the biochemical and cellular sequelae of seizure in the CNS, and point towards new avenues of research and potential therapeutic targets in epilepsy.
Animals↗