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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The proto-oncoprotein c-Rel is a member of the nuclear factor kappa B transcription factor family, which includes the p50 and p65 subunits of nuclear factor kappa B. We show here that c-Rel binds to kappa B sites as homodimers as well as heterodimers with p50. These homodimers and heterodimers show distinct DNA-binding specificities and affinities for various kappa B motifs. In particular, the c-Rel homodimer has a high affinity for interleukin-6 (IL-6) and beta interferon kappa B sites. In spite of its association with p50 in vitro, however, we found a lymphoid cell-specific nuclear factor in vivo that contains c-Rel but not p50 epitopes; this factor, termed IL-6 kappa B binding factor II, appears to contain the c-Rel homodimer and preferentially recognizes several IL-6 kappa B-related kappa B motifs. Although it has been previously shown that the IL-6 kappa B motif functions as a potent IL-1/tumor necrosis factor-responsive element in nonlymphoid cells, its activity was found to be repressed in lymphoid cells such as a Jurkat T-cell line. We also present evidence that IL-6 kappa B binding factor II functions as a repressor specific for IL-6 kappa B-related kappa B motifs in lymphoid cells.
Mol Cell Biol 1992 Apr
PMID:A lymphoid cell-specific nuclear factor containing c-Rel-like proteins preferentially interacts with interleukin-6 kappa B-related motifs whose activities are repressed in lymphoid cells. 137 88

Previously we have shown that nuclear extracts from mouse cells contain a heterogeneous group of polypeptides (p65, p80, p90, p100) which form distinct DNA-protein complexes on the 18 base-pair sequence element (termed Sal-box), which constitutes the murine rDNA transcription termination signal. These distinct proteins mediate cessation of RNA polymerase I (pol I) transcription elongation and release of the nascent RNA chains, indicating that they function as termination factor(s). Here, we report the biochemical analysis of the pol I-specific transcription termination factor TTFI. We show that the heterogeneity of TTFI is due to limited proteolysis of a larger, 130 kDa precursor protein (p130). The DNA-binding activity of p130 is strongly reduced as compared to the proteolytic derivatives, indicating that the DNA-binding domain is repressed within the full-length molecule. We have used limited proteolysis to purify and functionally characterize a TTFI core polypeptide (p50) which still specifically binds to the Sal-box target sequence and directs rDNA transcription termination. The equilibrium constant of purified p50 to bind specifically to DNA is 9 x 10(9) M-1. Additionally, we demonstrate that TTFI binds to DNA as a monomer and that binding induces DNA bending. This observation suggests that not only specific DNA-protein and protein-protein interactions but also conformational alterations of DNA may play a role in the termination process.
J Mol Biol 1992 Oct 05
PMID:Limited proteolysis unmasks specific DNA-binding of the murine RNA polymerase I-specific transcription termination factor TTFI. 140 80

Analysis of the p50 and p65 subunits of the NF-kappa B transcription factor complex has revealed that both proteins can interact with related DNA sequences through either homo- or heterodimer formation. In addition, the product of the proto-oncogene c-rel can bind to similar DNA motifs by itself or as a heterodimer with p50 or p65. However, these studies have used a limited number of known kappa B DNA motifs, and the question of the optimal DNA sequences preferred by each homodimer has not been addressed. Using purified recombinant p50, p65, and c-Rel proteins, optimal DNA-binding motifs were selected from a pool of random oligonucleotides. Alignment of the selected sequences allowed us to predict a consensus sequence for binding of the individual homodimeric Rel-related proteins, and DNA-protein binding analysis of the selected DNA sequences revealed sequence specificity of the proteins. Contrary to previous assumptions, we observed that p65 homodimers can interact with a subset of DNA sequences not recognized by p50 homodimers. Differential binding affinities were also obtained with p50- and c-Rel-selected sequences. Using either a p50- or p65-selected kappa B motif, which displayed differential binding with respect to the other protein, little to no binding was observed with the heterodimeric NF-kappa B complex. Similarly, in transfection experiments in which the selective kappa B binding sites were used to drive the expression of a chloramphenicol acetyltransferase reporter construct, the p65- and p50-selected motifs were activated only in the presence of p65 and p50/65 (a chimeric protein with the p50 DNA binding domain and p65 activation domain) expression vectors, respectively, and neither demonstrated a significant response to stimuli that induce NF-kappa B activity. These findings demonstrate that interaction of both subunits of the heterodimeric NF-kappa B complex with DNA is required for DNA binding and transcriptional activation and suggest that transcriptional activation mediated by the individual rel-related proteins will differ dramatically, depending on the specific kappa B motifs present.
Mol Cell Biol 1992 Oct
PMID:Selection of optimal kappa B/Rel DNA-binding motifs: interaction of both subunits of NF-kappa B with DNA is required for transcriptional activation. 140 30

Nuclear factor kappa B (NF-kappa B) modulates the expression of numerous genes via interaction with a specific DNA sequence termed the kappa B site. Its activity is modulated by a cytosolic inhibitor protein termed I kappa B, and its activation occurs in response to a variety of agents in a variety of cell types, most notably B and T lymphocytes. Data presented here show that an activity (designated complex I) that binds specifically to the kappa B site is induced in density-arrested Balb/c-3T3 mouse fibroblasts by platelet-derived growth factor (PDGF), a potent mitogen for these cells. Increased levels of complex I, as evaluated by electrophoretic mobility shift assays of nuclear extracts, were observed in cells treated for 1-4 h (but not 15 min) with the BB isoform of PDGF. 12-O-tetradecanoylphorbol 13-acetate (TPA) and the AA isoform of PDGF also stimulated this response and both isoforms, but not TPA, were effective in cells depleted of protein kinase C. Complex I most likely is authentic NF-kappa B, a p50-p65 heterodimer, or a closely related factor because it exhibited properties characteristic of those previously described for NF-kappa B including inducibility by deoxycholate and cycloheximide and sensitivity to I kappa B. A second kappa B binding activity (complex II), which apparently contained p50 homodimers, displayed limited induction by PDGF, whereas a third complex (complex III) migrated faster than but behaved similarly to complex I. These studies suggest that NF-kappa B or an NF-kappa B-like factor may participate in the expression of PDGF-inducible genes.
Mol Biol Cell 1992 Oct
PMID:Induction of NF-kappa B-like activity by platelet-derived growth factor in mouse fibroblasts. 142 70

The active nuclear form of the NF-kappa B transcription factor complex is composed of two DNA binding subunits, NF-kappa B p65 and NF-kappa B p50, both of which share extensive N-terminal sequence homology with the v-rel oncogene product. The NF-kappa B p65 subunit provides the transactivation activity in this complex and serves as an intracellular receptor for a cytoplasmic inhibitor of NF-kappa B, termed I kappa B. In contrast, NF-kappa B p50 alone fails to stimulate kappa B-directed transcription, and based on prior in vitro studies, is not directly regulated by I kappa B. To investigate the molecular basis for the critical regulatory interaction between NF-kappa B and I kappa B/MAD-3, a series of human NF-kappa B p65 mutants was identified that functionally segregated DNA binding, I kappa B-mediated inhibition, and I kappa B-induced nuclear exclusion of this transcription factor. Results from in vivo expression studies performed with these NF-kappa B p65 mutants revealed the following: 1) I kappa B/MAD-3 completely inhibits NF-kappa B p65-dependent transcriptional activation mediated through the human immunodeficiency virus type 1 kappa B enhancer in human T lymphocytes, 2) the binding of I kappa B/MAD-3 to NF-kappa B p65 is sufficient to retarget NF-kappa B p65 from the nucleus to the cytoplasm, 3) selective deletion of the functional nuclear localization signal present in the Rel homology domain of NF-kappa B p65 disrupts its ability to engage I kappa B/MAD-3, and 4) the unique C-terminus of NF-kappa B p65 attenuates its own nuclear localization and contains sequences that are required for I kappa B-mediated inhibition of NF-kappa B p65 DNA binding activity. Together, these findings suggest that the nuclear localization signal and transactivation domain of NF-kappa B p65 constitute a bipartite system that is critically involved in the inhibitory function of I kappa B/MAD-3. Unexpectedly, our in vivo studies also demonstrate that I kappa B/MAD-3 binds directly to NF-kappa B p50. This interaction is functional as it leads to retargeting of NF-kappa B p50 from the nucleus to the cytoplasm. However, no loss of DNA binding activity is observed, presumably reflecting the unique C-terminal domain that is distinct from that present in NF-kappa B p65.
Mol Biol Cell 1992 Dec
PMID:I kappa B/MAD-3 masks the nuclear localization signal of NF-kappa B p65 and requires the transactivation domain to inhibit NF-kappa B p65 DNA binding. 149 33

The cis-acting control elements of the interleukin-2 receptor alpha-chain (IL-2R alpha) gene contain a potent kappa B-like enhancer whose activity can be induced by various mitogenic stimuli. Recent cloning of the p50 and p65 subunits of the kappa B-binding protein NF-kappa B complex revealed a striking sequence homology of these proteins with the c-rel proto-oncogene product (c-Rel). On the basis of this homology, we examined the potential role of c-Rel in controlling IL-2R alpha transcription. We now demonstrate that the recombinant human c-Rel protein binds to the kappa B element in the IL-2R alpha promoter and results in alteration of the DNA structure in the adjacent downstream regulatory elements containing the CArG box and the GC box. We found that human c-Rel can activate transcription from the IL-2R alpha promoter, but not the kappa B-containing human immunodeficiency virus type 1 promoter, upon cotransfection into Jurkat T cells. Furthermore, truncation of the carboxyl terminus of c-Rel results in a c-Rel mutant (RelNA) that (i) localizes exclusively in the nucleus and (ii) acts in synergy with wild-type c-Rel in activating transcription from the kappa B site of the IL-2R alpha promoter. Finally, induction of surface IL-2R alpha expression coincides with the induced levels of endogenous c-Rel and induced c-Rel binding to the IL-2R alpha kappa B site. Our study identified c-Rel as one component of the Rel/NF-kappa B-family proteins involved in the kappa B-dependent activation of IL-2R alpha gene expression. Furthermore, our results suggest that a Re1NA-like cellular factor (e.g., NF-kappa B p50 or p49 subunit) acts in synergy with c-Re1 during T-cell activation.
Mol Cell Biol 1992 Sep
PMID:Kappa B site-dependent activation of the interleukin-2 receptor alpha-chain gene promoter by human c-Rel. 150 3

A Rel-related, mitogen-inducible, kappa B-binding protein has been cloned as an immediate-early activation gene of human peripheral blood T cells. The cDNA has an open reading frame of 900 amino acids capable of encoding a 97-kDa protein. This protein is most similar to the 105-kDa precursor polypeptide of p50-NF-kappa B. Like the 105-kDa precursor, it contains an amino-terminal Rel-related domain of about 300 amino acids and a carboxy-terminal domain containing six full cell cycle or ankyrin repeats. In vitro-translated proteins, truncated downstream of the Rel domain and excluding the repeats, bind kappa B sites. We refer to the kappa B-binding, truncated protein as p50B by analogy with p50-NF-kappa B and to the full-length protein as p97. p50B is able to form heteromeric kappa B-binding complexes with RelB, as well as with p65 and p50, the two subunits of NF-kappa B. Transient-transfection experiments in embryonal carcinoma cells demonstrate a functional cooperation between p50B and RelB or p65 in transactivation of a reporter plasmid dependent on a kappa B site. The data imply the existence of a complex family of NF-kappa B-like transcription factors.
Mol Cell Biol 1992 Feb
PMID:A novel mitogen-inducible gene product related to p50/p105-NF-kappa B participates in transactivation through a kappa B site. 153 Oct 86

When isolated from chick oviduct cytosol by antibody adsorption, the inactive progesterone receptor is associated with the two heat shock proteins, hsp90 and hsp70, plus three additional proteins termed p54, p50, and p23 according to their molecular weights. While their functions remain unknown, all of these receptor associated proteins are dissociated upon receptor activation in intact cells. To better understand the assembly and activation mechanisms of progesterone receptor complexes, we have developed a cell-free system for studying receptor interactions with hsp90 and hsp70 and have used this system to examine requirements for hsp90 binding to the receptor. Purified receptor, free of hsp90 and immobilized on an antibody affinity resin, will rebind hsp90 in rabbit reticulocyte lysate when several conditions are met. These include: (1) absence of progesterone, (2) elevated temperature (30 degrees C), (3) presence of ATP, and (4) presence of Mg2+. We have obtained maximal hsp90 binding to receptor when lysate is supplemented with 3 mM MgCl2 and an ATP regenerating system. ATP depletion of lysate by dialysis or ATPase addition blocks hsp90 binding to the receptor. When progesterone is added to pre-formed receptor complexes in reticulocyte lysate it promotes activation and the dissociation of hsp90. This process is also dependent upon ATP. Thus, both the assembly, and activation of the progesterone receptor can be accomplished in the reticulocyte lysate system.
J Steroid Biochem Mol Biol 1992 Mar
PMID:Composition, assembly and activation of the avian progesterone receptor. 156 3

The liver is one of the few adult tissues that has the capacity to regenerate following hepatectomy or toxic damage. In examining the early growth response during hepatic regeneration, we found that a highly induced immediate-early gene in regenerating liver encodes RL/IF-1 (regenerating liver inhibitory factor) and is the rat homolog of human MAD-3 and probably of chicken pp40. RL/IF-1 has I kappa B activity of broad specificity in that it inhibits the binding of p50-p65 NF-kappa B, c-Rel-p50, and RelB-p50, but not p50 homodimeric NF-kappa B, to kappa B sites. Like RL/IF-1, several members of the NF-kappa B and rel family of transcription factors are immediate-early genes in regenerating liver and mitogen-treated cells. We examined changes in kappa B site binding activity during liver regeneration and discovered a rapidly induced novel kappa B site-binding complex designated PHF [posthepatectomy factor(s)]. PHF is induced over 1,000-fold within minutes posthepatectomy in a protein synthesis-independent manner, with peak activity at 30 min, and is not induced by sham operation. PHF is distinct from p50-p65 NF-kappa B, which is present only in the inactive form in liver posthepatectomy. Although early PHF complexes do not interact strongly with anti-p50 antibodies, PHF complexes present later (3 to 5 h) posthepatectomy react strongly, suggesting that they contain a p50 NF-kappa B subunit. Unlike p50-p65 NF-kappa B, c-Rel-p50, and RelB-p50 complexes, PHF binding to kappa B sites is not inhibited by RL/IF-1. One role of RL/IF-1 in liver regeneration may be to inhibit p50-p65 NF-kappa B activity present in hepatic cells, allowing for the preferential binding of PHF to kappa B sites. Because PHF is induced immediately posthepatectomy in the absence of de novo protein synthesis, PHF could have a role in the regulation of liver-specific immediate-early genes in regenerating liver.
Mol Cell Biol 1992 Jun
PMID:Rapid induction in regenerating liver of RL/IF-1 (an I kappa B that inhibits NF-kappa B, RelB-p50, and c-Rel-p50) and PHF, a novel kappa B site-binding complex. 158 76

Mini-myoglobin, obtained by limited proteolysis of horse heart myoglobin (residues 32 to 139), represents a good model for testing the correlation between an exon and a protein domain. We have shown that ligand binding kinetics, spectral and folding features of mini-myoglobin are very similar to those of native myoglobin. In order to develop further the analysis of the structure-function relationship in this mini-protein, mini-globin was reconstituted with the heme moiety in which iron is replaced by cobalt. The Soret absorption spectra of oxy and deoxy cobaltous mini-myoglobin are very similar to those of cobaltous myoglobin derivatives; in addition. Co-mini-myoglobin binds oxygen reversibly with an n value approximately 1 and a p50 value of 45 to 50 mm Hg (the same as Co-myoglobin). Oxy Co-mini-myoglobin shows a well-resolved electron paramagnetic resonance (e.p.r.) spectrum typical of an oxygenated hemoprotein, while the spectrum of the deoxy derivative, although similar to that of deoxy Co-myoglobin, displays a lower resolution of the complex hyperfine structure. Moreover, photodissociation experiments on oxy Co-mini-myoglobin allow e.p.r. detection of an intermediate state, already observed in most hemoproteins and diagnostic for the interaction of bound oxygen with the distal histidine residue. Thus, reconstitution of mini-globin with cobalt protoprophyrin IX has provided, for the first time, a stable oxygenated complex that reflects a correct folding of the protein surrounding the heme pocket and possesses the functional behaviour typical of a hemoprotein.
J Mol Biol 1991 Dec 05
PMID:Mini-myoglobin. Electron paramagnetic resonance and reversible oxygenation of the cobalt derivative. 166 Sep 28


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