Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P05412 (c-Jun)
 11,453 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The addition of phorbol esters to U937 leukemic cells stimulates the phosphorylation of c-Jun on serines 63 and 73. To isolate the protein kinase which stimulates this phosphorylation, we have used heparin-Sepharose chromatography followed by affinity chromatography over glutathione-Sepharose beads bound with a fusion protein of glutathione S-transferase and amino acids 5-89 of c-Jun (GST-c-Jun). Using this procedure we purify a 67-kDa protein which is capable of phosphorylating GST-c-Jun as well as the complete c-Jun protein. By making mutations in serines 63 and 73 and then creating a fusion protein with GST (GST-c-Jun mut), we demonstrate that this protein kinase specifically phosphorylates these sites in the c-Jun amino terminus. Treatment of purified c-Jun amino-terminal protein kinase (cJAT-PK) with phosphatase 2A inhibits its ability to phosphorylate GST-c-Jun. This inactivated enzyme can be reactivated by phosphorylation with protein kinase C (PKC), although PKC is not capable of phosphorylating the GST-c-Jun substrate. Because v-Jun cannot be phosphorylated in vivo, we compared the ability of cJAT-PK to bind to GST-v-Jun or GST-c-Jun mut. The cJAT-PK bound 50-fold better to GST-c-Jun mut than GST-v-Jun suggesting that the delta domain which is missing in v-Jun plays a role in binding the cJAT-PK. These results suggest that there is a protein kinase cascade mediated by protein phosphatases and PKC which regulates c-Jun phosphorylation.
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PMID:Affinity-purified c-Jun amino-terminal protein kinase requires serine/threonine phosphorylation for activity. 132 19

c-Jun and its oncogenic counterpart v-Jun are completely conserved within the region from Ser-63 to Ser-73; these serines are sites for phorbol ester-inducible c-Jun phosphorylation. Using a U937 human leukemic cell line stably expressing v-Jun, we have demonstrated that phorbol esters stimulate the in vivo phosphorylation of c-Jun but not v-Jun. We developed an in vitro protein kinase assay to characterize the c-Jun protein kinase and to examine the determinants underlying this differential phosphorylation. Fusion proteins between glutathione S-transferase and the N terminus of c-Jun, v-Jun, or several c-Jun mutants were used as substrates. A c-Jun kinase activity was affinity-purified 5000-fold by using glutathione S-transferase-c-Jun-glutathione-Sepharose beads and was found to phosphorylate the N terminus of c-Jun but not v-Jun or c-Jun containing a 27-amino acid N-terminal deletion found in v-Jun. These effects were also observed in vivo as phorbol 12-myristate 13-acetate did not induce the phosphorylation of v-Jun or the c-Jun deletion mutant in U937 cell lines stably expressing these proteins. These findings indicate that the delta domain of c-Jun (amino acids 34-60), which is deleted in v-Jun, plays a critical role in regulating N-terminal c-Jun phosphorylation.
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PMID:Phorbol esters stimulate the phosphorylation of c-Jun but not v-Jun: regulation by the N-terminal delta domain. 160 42

The 86-kDa immediate-early 2 protein (IE2 86) of human cytomegalovirus is a powerful transactivator of homologous and heterologous promoters, including the human cytomegalovirus 1.2-kb RNA early promoter. Two potential mechanisms for gene activation by IE2 86 include interaction with cellular proteins and direct DNA binding. In this report, we show that the 1.2-kb RNA promoter contains a cis-acting AP-1 site, critical for its activation by IE2 86 in vivo, and that IE2 86, purified as a glutathione S-transferase-IE86 fusion protein, can interact with c-Jun and JunB. Additionally, by coimmunoprecipitation, we document that JunB and IE2 86 do associate in vivo. Further in vitro analysis reveals that Fos proteins are able to associate with glutathione S-transferase-IE86 only when present as a Jun-Fos heterodimer. With a set of IE2 86 mutants, we demonstrate that three independent regions of the IE2 86 interact in vitro with c-Jun, two of which are essential for activation of the 1.2-kb RNA promoter in vivo. We also show that IE2 86 can bind directly to this promoter through a sequence located just upstream of the AP-1 site between nucleotides -125 and -97. This discrete domain shares sequence homology with the cis-repression signal on the IE gene.
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PMID:The human cytomegalovirus IE2 86-kilodalton protein interacts with an early gene promoter via site-specific DNA binding and protein-protein associations. 766 55

Since the expression of glutathione S-transferase P-form (GST-P) has been suggested from in vitro studies to be partly regulated by the oncogene product, c-Jun and c-Fos, their distributions were compared in normal rat tissues and preneoplastic hepatic lesions induced by the Solt-Farber protocol. Immunohistochemically demonstrated GST-P protein was positively correlated with expression of both c-Jun and c-Fos in the epidermis of the skin and the smooth muscle of adult lung and with either c-Jun or c-Fos respectively in the bile ducts and bronchial epithelium. However, GST-P expression was also observed in proximal and distal straight segments of the kidney and other tissues negative for c-Jun and c-Fos and both c-Jun and c-Fos were present in the renal proximal and distal convoluted tubules, where GST-P was lacking. Thus, the localization of GST-P was in some cases clearly separable from those of c-Jun or c-Fos. GST-P was found to be focally expressed from an early stage of hepatocarcinogenesis, when c-Jun was not detectable. At later stages, this oncogene product was stained in 35.7% of GST-P-positive foci, with a clear relation to the degree of GST-P staining. Since GST-P is not always accompanied by appreciable c-Jun or c-Fos, these oncogene products are apparently not prerequisites for its expression. However, c-Jun may be partly responsible for maintaining high levels of GST-P in hepatic foci at later stages of hepatocarcinogenesis.
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PMID:Lack of correlated expression between the glutathione S-transferase P-form and the oncogene products c-Jun and c-Fos in rat tissues and preneoplastic hepatic foci. 769 15

c-Mil is the avian homologue of the mammalian serine/threonine kinase c-Raf-1. c-Mil/Raf is a mediator of signal transduction leading to gene expression via the c-Jun DNA-binding site, AP-1. Here we show that c-Mil immunopurified from MC29-virus-transformed quail fibroblasts phosphorylates c-Jun in vitro near its N terminus (Ser-63 and -73). Furthermore, the viral oncogene product Gag-Mil of the avian wild-type retrovirus MH2 phosphorylates c-Jun in vitro. A contribution by other known kinases phosphorylating c-Jun, such as the mitogen-activated protein kinases (MAPKs) and the c-Jun N-terminal kinases, was excluded by control reactions. c-Raf-1 and c-Jun directly interact in vitro as shown by various immobilized glutathione S-transferase-Raf fusion proteins which specify the cysteine-rich region of c-Mil/Raf as the major N-terminal binding site. An additional minor binding site is located in the C-terminal region. The biological relevance of these results is demonstrated by coimmunoprecipitation of c-Jun and c-Mil from 32P-labeled MC29- and MH2-transformed fibroblasts as well as normal quail embryo fibroblasts, whereby c-Jun was identified by tryptic phosphopeptide analysis. The complexed c-Jun exhibits a decreased electrophoretic mobility corresponding to a more highly phosphorylated state. Cell fractionation analyses indicate that the c-Mil/c-Jun complex is located in the cytoplasm. The data demonstrate that c-Jun can be a direct target of the protein kinase c-Mil/Raf, suggesting an alternative pathway, which leads to c-Jun phosphorylation independent of the MAPKs and MAPK-related proteins.
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PMID:Direct interaction and N-terminal phosphorylation of c-Jun by c-Mil/Raf. 787 94

Expression of the oncogene product c-Jun was examined and compared with that of class pi glutathione S-transferase (GST-II) during chemical hepatocarcinogenesis in female and male mice. A single i.p. injection of diethylnitrosamine (DEN) (10 mg/kg) was administered to B6C3F1 mice and livers were immunohistochemically investigated with anti-c-Jun and anti-GST-II antibodies at various time points thereafter. In females, almost all foci detected by hematoxylin and eosin staining were positive for c-Jun 3, 6, 9 and 11 months after the DEN administration. Seventy-one and 82% of c-Jun-positive foci at 9 and 11 months respectively, were also positive for GST-II, while this was the case for only 15% at 6 months. In males almost all foci were also positive for c-Jun at 3 and 6 months, but 23% of foci were negative at 9 months. Unlike the foci in females, 96 and 79% of those in males expressing c-Jun were negative for GST-II at 6 and 9 months respectively. Both GST-II expression in foci of females and its lack in those of males were highly correlated with c-Jun expression. Furthermore, single cells expressing c-Jun were also observed in both sexes at 3 months and thereafter, but not at 2 or 4 weeks. Alterations in the numbers of c-Jun-positive single cells, minifoci and foci followed sequentially revealed the number of such single cells to decrease, while foci increased, the sum being relatively constant. On the other hand, while a large number of GST-II-positive single cells were detected in female livers 2 and 4 weeks after the DEN administration, they markedly decreased thereafter, suggesting that the majority were unlikely to give rise to foci. Thus, c-Jun may be a better positive marker not only for preneoplastic foci, but also putative precursor single cells in both female and male mice and therefore be useful for analysis of hepatocarcinogenic processes.
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PMID:c-Jun expression in single cells and preneoplastic foci induced by diethylnitrosamine in B6C3F1 mice: comparison with the expression of pi-class glutathione S-transferase. 792 77

Human proenkephalin gene transcription is transactivated by human T-cell leukemia virus type I (HTLV-I) Tax in human Jurkat T lymphocytes. This transactivation was further enhanced in Jurkat cells treated with concanavalin A, cyclic AMP, or 12-O-tetradecanoylphorbol-13-acetate. Deletion and cis-element transfer analyses of the human proenkephalin promoter identified a cyclic AMP-responsive AP-1 element (-92 to -86) as both necessary and sufficient to confer Tax-dependent transactivation. Different AP-1 or cyclic AMP-responsive element-binding protein (CREB)/activating transcription factor (ATF) proteins which bind this element were expressed in murine teratocarcinoma F9 cells to identify those capable of mediating Tax-dependent transactivation of human proenkephalin gene transcription. Although CREB, c-Fos, c-Jun, and JunD did not have significant effects, JunB inhibited the Tax-dependent transactivation. In contrast, ATF3 dramatically induced Tax-dependent transactivation, which was further enhanced by protein kinase A. Electrophoretic mobility shift assays with recombinant fusion proteins expressed and purified from bacteria indicate that the DNA-binding activity of ATF3 is also dramatically enhanced by Tax. Chimeric fusion proteins consisting of the DNA-binding domain of the yeast transcription factor Gal4 and the amino-terminal domain (residues 1 to 66) of ATF3 were able to mediate Tax-dependent transactivation of a Gal4-responsive promoter, which suggests a direct involvement of this region of ATF3. Recombinant fusion proteins of glutathione S-transferase with either the amino- or carboxy-terminal (residues 139 to 181) domain of ATF3 were able to specifically interact with Tax. Furthermore, specific antisera directed against Tax coimmunoprecipitated ATF3 only in the presence of Tax.
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PMID:Novel interactions between human T-cell leukemia virus type I Tax and activating transcription factor 3 at a cyclic AMP-responsive element. 800 91

Association of the human c-Jun and c-Fos proteins depends upon interactions involving their leucine zipper domains. We are interested in elucidating the tertiary structure of the Jun and Fos leucine zipper domains with a view to understanding the precise intermolecular interactions which govern the affinity and specificity of interaction in these proteins, which have the unusual capacity to form either homodimeric or heterodimeric zipper pairs. With this goal in mind, we have developed a bacterial expression system for the efficient production of both unlabelled and isotopically labelled c-Jun leucine zipper domain. A synthetic junLZ gene was created by annealing, ligation, and polymerase-chain-reaction amplification of overlapping synthetic oligonucleotides which comprised 132 bp of coding sequence encompassing residues Arg276-Asn314 of c-Jun plus a total of five engineered non-native residues at the N- and C-termini. The junLZ gene was cloned into the pGEX-2T vector from which recombinant c-Jun leucine zipper domain (rJunLZ; 46 residues, 5.1 kDa) was overexpressed (approximately 15% total cell protein) in Escherichia coli as a fusion protein of 31.4 kDa, consisting of rJunLZ fused to the carboxy-terminal portion of Schistosoma japonicum glutathione S-transferase. Two markedly different expression strategies have been devised which allow purification of rJunLZ from the soluble or inclusion-body fraction of induced cells. We have used these strategies to produce unlabelled and uniformly 15N-labelled rJunLZ for NMR studies which, in combination with circular dichroic measurements, reveal that rJunLZ most likely forms a symmetric coiled-coil of parallel alpha-helices. We also present 15N-NMR chemical shift assignments for the backbone and sidechain amide nitrogens of rJunLZ, which should assist in determination of a high-resolution structure of the homodimeric Jun leucine zipper using heteronuclear three-dimensional NMR spectroscopy.
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PMID:Cloning, expression, and spectroscopic studies of the Jun leucine zipper domain. 811 39

Induction of mouse glutathione S-transferase (GST) Ya gene expression by a variety of chemical agents is mediated by a regulatory element, EpRE, composed of an Ets and two adjacent AP-1-like binding sites. In this report we present evidence that the basal and inducible activity of EpRE is mediated by AP-1 transcription factor and that the cooperative interaction between AP-1 and an Ets protein contributes to enhance the EpRE inducibility. We also show that EpRE, similar to a single AP-1 site, when ligated to GST Ya gene promoter, is transactivated by c-Fos/c-Jun or c-Fos/Jun-B heterodimer and that c-Jun/c-Jun homodimer is an activator of an AP-1 site only in the context of collagenase gene promoter.
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PMID:Cooperative interaction between Ets and AP-1 transcription factors regulates induction of glutathione S-transferase Ya gene expression. 816 97

Elevated levels of the human pi class glutathione S-transferase (GSTP1-1) have been implicated in the development of antineoplastic drug resistance. Using GSTP1 promoter deletion constructs we have shown that enhanced GSTP1 transcription (up to 18-fold) is the predominant mechanism responsible for increased GSTP1-1 levels in a multidrug resistant derivative (VCREMS) of the human mammary carcinoma cell line MCF7. Furthermore, disruption of a putative AP-1 response element within the GSTP1 promoter (nucleotides -69 to -63) abrogated GSTP1 transcription in both cell lines. In addition, band shift assays demonstrated binding of a VCREMS nuclear complex to the promoter region C1 (-73 to -54) which could be competed for by a DNA fragment containing a known AP-1 binding site from the human collagenase promoter. However, no such competition was observed for the major MCF7 C1 complex. The role of a Fos-Jun-like complex in regulating GSTP1 transcription in VCREMS cells was further emphasized by the introduction of point mutations within the C1 region which were known to inhibit AP-1 binding and the interaction of antisera raised against human c-Jun and c-Fos with the major C1 complex in VCREMS cells. These studies therefore highlight cell-specific differences in the binding pattern of Jun and Fos proteins to the GSTP1 promoter which are likely to play an important role in regulating transcriptional activation of the GSTP1 gene in drug-resistant breast cancer cells.
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PMID:Involvement of Jun and Fos proteins in regulating transcriptional activation of the human pi class glutathione S-transferase gene in multidrug-resistant MCF7 breast cancer cells. 820 48


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