C A Rosen

Human Genome Sciences, Maryland, United States

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Publications (79)914.32 Total impact

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    ABSTRACT: Our laboratory previously described the independent isolation of the fibroblast growth factor 4 (FGF-4) gene by NIH3T3 transformation assay using DNA from a patient with CML leukemia (Lucas et al., 1994). The FGF-4 gene was truncated by DNA rearrangement with a novel gene named GRS. In this manuscript we describe isolation of GRS cDNA and show by sequence comparison that GRS is a novel member of the Bcl-2 gene family. Northern analysis shows expression of the gene in normal human tissue to be largely restricted to the hematopoietic compartment. Analysis of the pattern of gene expression in cancer cell lines demonstrates GRS is expressed in hematopoietic malignancies and in melanoma. The chromosomal location of GRS has also been determined. The gene is positioned on chromosome 15 within bands q24-25.
    Oncogene 03/1997; 14(8):997-1001. · 8.56 Impact Factor
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    ABSTRACT: The therapeutic significance of recombinant human DNase I in treating the patients with cystic fibrosis has risen our interests in identifying other human DNase I-like enzymes to study their biological significance. Here we described our work of cloning and characterization of a novel gene, which encodes a human protein homologous to human DNase I. A full length cDNA clone of this gene consists of 1290 bp, encoding a polypeptide of 306 amino acids. The deduced amino acid sequence of this novel human DNase (nhDNase) is 45% identical to that of human DNase I. Among sixteen human tissues examined by Northern Blot, high level expression of nhDNase was found in human liver and spleen. Recombinant protein of nhDNase was produced in a Baculovirus expression system and purified by chromatography and reverse-phase HPLC. Purified recombinant nhDNase migrated as a single band of about 33 kD molecular weight analyzed by SDS-PAGE. The DNase activity of nhDNase was demonstrated by assay of hydrolysis of S.S.DNA. Its activity was dependent upon the presence of divalent metal irons, calcium and magnesium. However, unlike bovine pancreas DNase I, nhDNase was not inhibited by G-actin of bovine muscle, which indicates the physiological significance of this enzyme in clinical implication.
    Biochemical and Biophysical Research Communications 03/1997; 231(2):499-504. · 2.28 Impact Factor
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    ABSTRACT: NF-kappaB is a pleiotropic transcriptional activator originally identified by its ability to regulate immunoglogulin kappa light chain expression. Purification of this DNA-binding complex demonstrated that NF-kappaB is a heterodimer composed of two subunits, NFKB1 and RelA. Previous studies have shown that truncated versions of these proteins could be expressed and purified from bacterial cells. In the present study, we utilize a baculovirus expression vector system (BEVS) to overexpress each subunit independently to produce homodimers or together to reconstitute functional NF-kappaB. These proteins can be enriched to >70% homogeneity on a kappaB-agarose DNA- affinity column. The purified proteins are active in DNA binding as measured by electrophoretic mobility shift assays. Finally, transcriptional activation of these recombinant proteins can be measured by their ability to activate a kappaB-CAT reporter plasmid in transiently transfected/infected SF-9 cells. Thus, BEVS provides a method for production of full-length, transcriptionally active NF-kappaB proteins.
    Protein Expression and Purification 02/1997; 9(1):40-8. · 1.43 Impact Factor
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    ABSTRACT: Stanniocalcin (STC) is a glycoprotein hormone first identified in bony fishes where it counteracts hypercalcemia by inhibiting gill calcium uptake and stimulating renal inorganic phosphate (Pi) reabsorption. Human STC (hSTC) has recently been cloned and sequenced and is highly homologous to the fish hormone at the amino acid level. The objective of this study was to examine the possible effects of hSTC on electrolyte homeostasis and renal function in the rat. Recombinant hSTC was expressed in bacteria and purified by metal-ion affinity chromatography and reverse-phase high performance liquid chromatography. Anesthetized animals were given bolus infusions of 1, 5, or 10 nmol hSTC per kilogram of body weight. Control animals received solvent alone. The most effective dosage was 5 nmol/kg, which caused significant reductions in both absolute and fractional phosphate excretion in comparison with control rats. The hSTC had no effect on the renal excretion of other ions, the glomerular filtration rate, renal blood flow, blood pressure, or plasma electrolytes (Na+, K+, Ca2+, Pi, Mg/+). The maximum effect of hSTC on phosphate excretion was observed 60-80 minutes postinjection. Lesser effects were obtained with higher and lower dosages of hormone. When renal cortical brush-border membrane vesicles were isolated from control and hormone-treated animals 80 minutes postinjection, the rate of Na+/Pi cotransport was found to be 40% higher in vesicles from hormone-treated animals (p < 0.01; 5 nmol hSTC/kg). Together, the renal clearance and membrane vesicle data indicate that hSTC participates in the renal regulation of Pi homeostasis in mammals.
    Journal of Bone and Mineral Research 02/1997; 12(2):165-71. · 6.13 Impact Factor
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    ABSTRACT: The ubiquitous transcription factor NF-kappaB is an essential component in signal transduction pathways, in inflammation, and in the immune response. NF-kappaB is maintained in an inactive state in the cytoplasm by protein-protein interaction with IkappaBalpha. Upon stimulation, rapid degradation of IkappaBalpha allows nuclear translocation of NF-kappaB. To study the importance of IkappaBalpha in signal transduction, IkappaBalpha-deficient mice were derived by gene targeting. Cultured fibroblasts derived from IkappaBalpha-deficient embryos exhibit levels of NF-kappaB1, NF-kappaB2, RelA, c-Rel, and IkappaBbeta similar to those of wild-type fibroblasts. A failure to increase nuclear levels of NF-kappaB indicates that cytoplasmic retention of NF-kappaB may be compensated for by other IkappaB proteins. Treatment of wild-type cells with tumor necrosis factor alpha (TNF-alpha) resulted in rapid, transient nuclear localization of NF-kappaB. IkappaBalpha-deficient fibroblasts are also TNF-alpha responsive, but nuclear localization of NF-kappaB is prolonged, thus demonstrating that a major irreplaceable function Of IkappaBalpha is termination of the NF-kappaB response. Consistent with these observations, and with IkappaBalpha and NF-kappaB's role in regulating inflammatory and immune responses, is the normal development Of IkappaBalpha-deficient mice. However, growth ceases 3 days after birth and death usually occurs at 7 to 10 days of age. An increased percentage of monocytes/macrophages was detected in spleen cells taken from 5-, 7-, and 9-day-old pups. Death is accompanied by severe widespread dermatitis and increased levels of TNF-alpha mRNA in the skin.
    Molecular and Cellular Biology 06/1996; 16(5):2341-9. · 5.37 Impact Factor
  • P A Moore, C A Rosen, K C Carter
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    ABSTRACT: This report describes the localization of the human FKBP12-rapamycin-associated protein (FRAP) gene to human chromosome 1p36 using fluorescence in situ hybridization. This protein is the binding site for rapamycin and FK506, two potent immunosuppressive drugs. 12 refs., 1 fig.
    Genomics 05/1996; 33(2):331-2. · 3.01 Impact Factor
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    ABSTRACT: We have isolated a human cDNA clone encoding the mammalian homolog of stanniocalcin (STC), a calcium- and phosphate-regulating hormone that was first described in fishes where it functions in preventing hypercalcemia. STC has a unique amino acid sequence and, until now, has remained one of the few polypeptide hormones never described in higher vertebrates. Human STC (hSTC) was found to be 247 amino acids long and to share 73% amino acid sequence similarity with fish STC. Polyclonal antibodies to recombinant hSTC localized to a distinct cell type in the nephron tubule, suggesting kidney as a possible site of synthesis. Recombinant hSTC inhibited the gill transport of calcium when administered to fish and stimulated renal phosphate reabsorption in the rat. The evidence suggests that mammalian STC, like its piscine counterpart, is a regulator of mineral homeostasis.
    Proceedings of the National Academy of Sciences 04/1996; 93(5):1792-6. · 9.81 Impact Factor
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    ABSTRACT: Transcriptional adaptor proteins are required for full function of higher eukaryotic acidic activators in the yeast Saccharomyces cerevisiae, suggesting that this pathway of activation is evolutionarily conserved. Consistent with this view, we have identified possible human homologs of yeast ADA2 (yADA2) and yeast GCN5 (yGCN5), components of a putative adaptor complex. While there is overall sequence similarity between the yeast and human proteins, perhaps more significant is conservation of key sequence features with other known adaptors. We show several functional similarities between the human and yeast adaptors. First, as shown for yADA2 and yGCN5, human ADA2 (hADA2) and human GCN5 (hGCN5) interacted in vivo in a yeast two-hybrid assay. Moreover, hGCN5 interacted with yADA2 in this assay, suggesting that the human proteins form similar complexes. Second, both yADA2 and hADA2 contain cryptic activation domains. Third, hGCN5 and yGCN5 had similar stabilizing effects on yADA2 in vivo. Furthermore, the region of yADA2 that interacted with yGCN5 mapped to the amino terminus of yADA2, which is highly conserved in hADA2. Most striking, is the behavior of the human proteins in human cells. First, GAL4-hADA2 activated transcription in HeLa cells, and second, either hADA2 or hGCN5 augmented GAL4-VP16 activation. These data indicated that the human proteins correspond to functional homologs of the yeast adaptors, suggesting that these cofactors play a key role in transcriptional activation.
    Molecular and Cellular Biology 03/1996; 16(2):593-602. · 5.37 Impact Factor
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    ABSTRACT: TFIIA is thought to play an important role in transcriptional regulation in higher eukaryotes, but its precise function is unclear. A human cDNA encoding a protein with 45% identity to the small subunit of yeast TFIIA has been isolated. TFIIA activity could be reconstituted by the mixing of recombinant large (alpha beta) and small (gamma) subunits. TFIIA-depleted HeLa nuclear extracts were used to demonstrate that TFIIA is essential for basal and activated transcription by several distinct classes of activators. Recombinant TFIIA functioned in transcriptional activation whether expressed as a dimer (alpha beta+gamma) or as a trimer (alpha+beta+gamma), which closely resembles the native form. Yeast TFIIA also functioned in transcriptional activation, and the human gamma subunit was functionally interchangeable with TOA2, its yeast homolog. Recombinant TFIIA mediated the stimulation of TFIID binding to the TATA region and downstream promoter sequences by the Zta transcriptional activator. Significantly, we found that TFIIA bound directly to Zta in an activation domain-dependent manner. One consequence of the TFIIA-mediated interaction between Zta and TFIID was the formation of a promoter-bound complex resistant to TATA oligonucleotide competition. These results demonstrate that TFIIA is an evolutionarily conserved general factor critical for activator-regulated transcription.
    Genes & Development 11/1994; 8(19):2324-35. · 12.44 Impact Factor
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    ABSTRACT: Hereditary nonpolyposis colorectal cancer (HNPCC) is one of man's commonest hereditary diseases. Several studies have implicated a defect in DNA mismatch repair in the pathogenesis of this disease. In particular, hMSH2 and hMLH1 homologues of the bacterial DNA mismatch repair genes mutS and mutL, respectively, were shown to be mutated in a subset of HNPCC cases. Here we report the nucleotide sequence, chromosome localization and mutational analysis of hPMS1 and hPMS2, two additional homologues of the prokaryotic mutL gene. Both hPMS1 and hPMS2 were found to be mutated in the germline of HNPCC patients. This doubles the number of genes implicated in HNPCC and may help explain the relatively high incidence of this disease.
    Nature 10/1994; 371(6492):75-80. · 38.60 Impact Factor
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    ABSTRACT: HEREDITARY nonpolyposis colorectal cancer (HNPCC) is one of man's commonest hereditary diseases1. Several studies have implicated a defect in DNA mismatch repair in the pathogenesis of this disease2-8. In particular, hMSH2 and hMLHl homologues of the bacterial DNA mismatch repair genes mutS and mutL, respectively, were shown to be mutated in a subset of HNPCC cases9-16. Here we report the nucleotide sequence, chromosome localization and mutational analysis of hPMSl and hPMS2, two additional homologues of the prokaryotic mutL gene. Both hPMSl and hPMS2 were found to be mutated in the germline of HNPCC patients. This doubles the number of genes implicated in HNPCC and may help explain the relatively high incidence of this disease.
    Nature 09/1994; 371(6492):75-80. · 38.60 Impact Factor
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    ABSTRACT: Transcriptional activation of the IL-8 gene by several inflammatory mediators, including the cytokines IL-1 and TNF-alpha, is mediated through sequences located between nucleotide -94 and -71 of the IL-8 promoter. Because adjacent binding sites for the inducible transcription factors NF-kappa B and NF-IL-6 are located within this region, we examined the functional interaction of these two transcription factor families in IL-8 gene regulation. Maximal transcriptional activation by PMA in Jurkat T lymphocytes was shown to require intact binding sites for both NF-kappa B and NF-IL-6. Electrophoretic mobility shift analysis indicates that NF-IL-6, as well as other related members of this family, bind specifically to the NF-IL-6 site in the IL-8 promoter. In addition, NF-kappa B p65 (RelA), but not NF-kappa B p50 (NFKB1), binds specifically to the NF-kappa B site. When incubated together, RelA and NF-IL-6/C/EBP form a ternary complex with this region of the IL-8 promoter; this binding is dependent on intact binding sites for both NF-IL-6 and RelA. Transient cotransfection analyses indicate that the cooperative association of NF-IL-6 and RelA with the IL-8 promoter results in synergistic transcriptional activation. Mutational analyses of RelA demonstrate that the C-terminal transactivation domain and the DNA binding domain are required for synergistic activation with NF-IL-6. In addition, overexpression of the NF-kappa B inhibitor molecule, I kappa B, abolished the RelA- and RelA/NF-IL-6-dependent synergistic activation. These data demonstrate that RelA and members of the C/EBP/NF-IL-6 family can functionally cooperate in transcriptional activation of the IL-8 gene and suggest a common mechanism for inducible regulation of cytokine gene expression.
    The Journal of Immunology 08/1994; 153(1):153-64. · 5.52 Impact Factor
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    ABSTRACT: Exposure of monocytic cells to bacterial lipopolysaccharide (LPS) activates the NF-kappa B/Rel family of proteins and leads to the rapid induction of inflammatory gene products, including tissue factor (TF). TF is the primary cellular initiator of the coagulation protease cascades. Here we report the characterization of a nuclear complex from human monocytic cells that bound to a kappa B-like site, 5'-CGGAGTTTCC-3', in the 5'-flanking region of the human TF gene. This nuclear complex was activated by LPS with kinetics that preceded induction of the TF gene. In vitro binding studies demonstrated that the TF site bound translated c-Rel and p65 homodimers but not p50/p65 heterodimers or p50 homodimers. Base-pair substitutions in the TF site indicated that the presence of a cytosine at position 1 precluded binding of NF-kappa B. In fact, under low-ionic-strength conditions, the TF complex did not migrate with translated p50/p65 dimers but instead comigrated with c-Rel/p65 dimers. Antibodies against the NF-kappa B and Rel proteins and UV cross-linking studies revealed the presence of c-Rel and p65 and the absence of p50 in the TF complex and further showed that c-Rel/p65 heterodimers selectively bound to the TF kappa B-like site. Functional studies indicated that the TF site conferred LPS inducibility on a heterologous promoter and was transactivated by c-Rel or p65. Taken together, our results demonstrated that binding of c-Rel/p65 heterodimers to a novel kappa B-like site mediated LPS induction of TF gene expression in monocytic cells.
    Molecular and Cellular Biology 07/1994; 14(6):3772-81. · 5.37 Impact Factor
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    ABSTRACT: Some cases of hereditary nonpolyposis colorectal cancer (HNPCC) are due to alterations in a mutS-related mismatch repair gene. A search of a large database of expressed sequence tags derived from random complementary DNA clones revealed three additional human mismatch repair genes, all related to the bacterial mutL gene. One of these genes (hMLH1) resides on chromosome 3p21, within 1 centimorgan of markers previously linked to cancer susceptibility in HNPCC kindreds. Mutations of hMLH1 that would disrupt the gene product were identified in such kindreds, demonstrating that this gene is responsible for the disease. These results suggest that defects in any of several mismatch repair genes can cause HNPCC.
    Science 04/1994; 263(5153):1625-9. · 31.03 Impact Factor
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    ABSTRACT: The NF-kappa B transcription factor, composed of two proteins, p50 and p65, is a pleiotropic activator that participates in the induction of a wide variety of cellular genes. Various cell adhesion molecules have NF-kappa B binding sites and may play an important role in inflammatory response, tumorigenicity, and metastasis. In an earlier study, we demonstrated that adhesion of diverse transformed cells was blocked by antisense inhibition of the p65 subunit of NF-kappa B. Since cell-substratum interactions play an important role in tumorigenicity, we reasoned that antisense p65 could inhibit tumorigenicity. In diverse transformed cell lines, phosphorothioate antisense oligonucleotides to p65 inhibited in vitro growth, reduced soft-agar colony formation, and eliminated the ability of cells to adhere to an extracellular matrix. Stable transfectants of a fibrosarcoma cell line expressing dexamethasone-inducible antisense RNA to p65 showed inhibition of in vitro growth and in vivo tumor development. In response to inducible expression of antisense RNA, a pronounced tumor regression was seen in nude mice. The administration of antisense but not sense p65 oligonucleotides caused a pronounced inhibition of tumorigenicity in nude mice injected with diverse tumor-derived cell lines. Inhibitors of NF-kappa B function may thus be useful in the treatment of cancer.
    Proceedings of the National Academy of Sciences 12/1993; 90(21):9901-5. · 9.81 Impact Factor
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    C Kunsch, C A Rosen
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    ABSTRACT: Interleukin-8 (IL-8), a chemotactic cytokine for T lymphocytes and neutrophils, is induced in several cell types by a variety of stimuli including the inflammatory cytokines IL-1 and tumor necrosis factor alpha TNF-alpha. Several cis elements, including a binding site for the inducible transcription factor NF-kappa B, have been identified in the regulatory region of the IL-8 gene. We have examined the ability of various NF-kappa B subunits to bind to, and activate transcription from, the IL-8 promoter. A nuclear complex was induced in phorbol myristate acetate-treated Jurkat T cells which bound specifically to the kappa B site of the IL-8 promoter and was inhibited by addition of purified I kappa B alpha to the reaction mixture. Only antibody to RelA (p65), but not to NFKB1 (p50), NFKB2 (p50B), c-Rel, or RelB was able to abolish binding, suggesting that RelA is a major component in these kappa B binding complexes. Gel mobility shift analysis with in vitro-translated and purified proteins indicated that whereas the kappa B element in the human immunodeficiency virus type 1 long terminal repeat bound to all members of the kappa B/Rel family examined, the IL-8 kappa B site bound only to RelA and to c-Rel and NFKB2 homodimers, but not to NFKB1 homodimers or heterodimers of NFKB1-RelA. Transient transfection analysis demonstrated a kappa B-dependent expression of the IL-8 promoter in a human fibrosarcoma cell line (8387) and in Jurkat T lymphocytes. Cotransfection with various NF-kappa B subunits indicated that RelA and c-Rel, but neither NFKB1 nor heterodimeric NFKB1-RelA, was able to activate transcription from the IL-8 promoter. Furthermore, cotransfection of NFKB1 and RelA, although able to support activation from the human immunodeficiency virus type 1 long terminal repeat, failed to activate expression from the IL-8 promoter. Antisense oligonucleotides to RelA, but not NFKB1, inhibited phorbol myristate acetate-induced IL-8 production in Jurkat T lymphocytes. These data demonstrate the differential ability of members of the kappa B/Rel family to bind to, and activate transcription from, the IL-8 promoter. Furthermore, while providing a novel example of a kappa B-regulated promoter in which the classical NF-kappa B complex is unable to activate transcription from the kappa B element, these data provide direct evidence for the role of RelA in regulation of IL-8 gene expression.
    Molecular and Cellular Biology 11/1993; 13(10):6137-46. · 5.37 Impact Factor
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    ABSTRACT: Oxidative stress and expression of the vascular cell adhesion molecule-1 (VCAM-1) on vascular endothelial cells are early features in the pathogenesis of atherosclerosis and other inflammatory diseases. Regulation of VCAM-1 gene expression may be coupled to oxidative stress through specific reduction-oxidation (redox) sensitive transcriptional or posttranscriptional regulatory factors. In cultured human umbilical vein endothelial (HUVE) cells, the cytokine interleukin 1 beta (IL-1 beta) activated VCAM-1 gene expression through a mechanism that was repressed approximately 90% by the antioxidants pyrrolidine dithiocarbamate (PDTC) and N-acetylcysteine (NAC). Furthermore, PDTC selectively inhibited the induction of VCAM-1, but not intercellular adhesion molecule-1 (ICAM-1), mRNA and protein accumulation by the cytokine tumor necrosis factor-alpha (TNF alpha) as well as the noncytokines bacterial endotoxin lipopolysaccharide (LPS) and double-stranded RNA, poly(I:C) (PIC). PDTC also markedly attenuated TNF alpha induction of VCAM-1-mediated cellular adhesion. In a distinct pattern, PDTC partially inhibited E-selectin gene expression in response to TNF alpha but not to LPS, IL-1 beta, or PIC. TNF alpha and LPS-mediated transcriptional activation of the human VCAM-1 promoter through NF-kappa B-like DNA enhancer elements and associated NF-kappa B-like DNA binding proteins was inhibited by PDTC. These studies suggest a molecular linkage between an antioxidant sensitive transcriptional regulatory mechanism and VCAM-1 gene expression that expands on the notion of oxidative stress as an important regulatory signal in the pathogenesis of atherosclerosis.
    Journal of Clinical Investigation 11/1993; 92(4):1866-74. · 12.81 Impact Factor
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    ABSTRACT: NF-kappa B is a pleiotropic regulator of a variety of genes implicated in the cellular response to injury. This function has been attributed to the coordinated binding of subunits of NF-kappa B to distinct regions of the promoter elements of numerous genes, including cytokines, growth factor receptors, and adhesion molecules. Antisense phosphorothioate oligonucleotides to the p50 and p65 subunits of the NF-kappa B complex were used to define the physiologic role of this transcription factor in resting and stimulated granulocytes. A reduction in the expression of p65 was produced by treatment with the phosphorothioate antisense oligodeoxynucleotide. This reduction was accompanied by rapid changes in the cellular adhesion of dimethyl sulfoxide-differentiated HL-60 leukemia cells stimulated by 12-O-tetradecanoylphorbol 13-acetate (TPA). These effects were characterized by a marked reduction in CD11b integrin expression on the surface of treated cells. Furthermore, the p65 antisense oligomer effectively abolished an upregulation of CD11b that was produced by formyl-met-leu-phe and TPA. However, the p65 antisense phosphorothioate oligodeoxynucleotide had no significant effect on the production of reactive oxygen intermediates or on phagocytosis by these cells. These findings indicate that antisense oligomers to p65 can be used to define the role of NF-kappa B in the activation pathways of neutrophils.
    Blood 08/1993; 82(2):625-32. · 9.78 Impact Factor
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    ABSTRACT: The subunits of NF-kappa B, NFKB1 (formerly p50) and RelA (formerly p65), belong to a growing family of transcription factors that share extensive similarity to the c-rel proto-oncogene product. The homology extends over a highly conserved stretch of approximately 300 amino acids termed the Rel homology domain (RHD). This region has been shown to be involved in both multimerization (homo- and heterodimerization) and DNA binding. It is now generally accepted that homodimers of either subunit are capable of binding DNA that contains a kappa B site originally identified in the immunoglobulin enhancer. Recent studies have demonstrated that the individual subunits of the NF-kappa B transcription factor complex can be distinguished by their ability to bind distinct DNA sequence motifs. By using NFKB1 and RelA subunit fusion proteins, different regions within the RHD were found to confer DNA-binding and multimerization functions. A fusion protein that contains 34 N-terminal amino acids of NFKB1 and 264 amino acids of RelA displayed preferential binding to an NFKB1-selective DNA motif while dimerizing with the characteristics of RelA. Within the NFKB1 portion of this fusion protein, a single amino acid change of His to Arg altered the DNA-binding specificity to favor interaction with the RelA-selective DNA motif. Furthermore, substitution of four amino acids from NFKB1 into RelA was able to alter the DNA-binding specificity of the RelA protein to favor interaction with the NFKB1-selective site. Taken together, these findings demonstrate the presence of a distinct subdomain within the RHD involved in conferring the DNA-binding specificity of the Rel family of proteins.
    Molecular and Cellular Biology 08/1993; 13(7):3850-9. · 5.37 Impact Factor
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    ABSTRACT: The p50 and p65 subunits of NF-kappa B represent two members of a gene family that shares considerable homology to the rel oncogene. Proteins encoded by these genes form homo- and heterodimers which recognize a common DNA sequence motif. Recent data have suggested that homodimers of individual subunits of NF-kappa B can selectively activate gene expression in vitro. To explore this possibility in a more physiological manner, murine embryonic stem (ES) cells were treated with phosphorothio antisense oligonucleotides to either p50 or p65. Within 5 h after exposure to phosphorothio antisense p65 oligonucleotides, cells exhibited dramatic alterations in adhesion properties. Similar findings were obtained in a stable cell line that expressed a dexamethasone-inducible antisense mRNA to p65. Although antisense oligonucleotides raised against both p50 and p65 elicited a significant reduction in their respective mRNAs, only the cells treated with antisense p50 maintained a normal morphology. However, 6 days following removal of leukemia-inhibiting factor, a growth factor which suppresses embryonic stem cell differentiation, adhesion properties of cells treated with the antisense p50 oligonucleotides were markedly affected. The ability of the individual antisense oligonucleotides to elicit differential effects on cell adhesion, a property dependent upon the stage of differentiation, suggests that the p50 and p65 subunits of NF-kappa B regulate gene expression either as homodimers or as heterodimers with other rel family members. Furthermore, the finding that reduction in p65 expression alone had profound effects on cell adhesion properties indicates that p65 plays an important role in nonstimulated cells and cannot exist solely complexed with the cytosolic inhibitory protein I kappa B.
    Molecular and Cellular Biology 07/1993; 13(6):3802-10. · 5.37 Impact Factor

Publication Stats

10k Citations
914.32 Total Impact Points

Institutions

  • 1993
    • Human Genome Sciences
      Maryland, United States
  • 1988–1993
    • Roche Institute of Molecular Biology
      Nutley, New Jersey, United States
  • 1992
    • University of Massachusetts Medical School
      • Department of Cell Biology
      Worcester, MA, United States
  • 1986–1988
    • Dana-Farber Cancer Institute
      • Department of Cancer Immunology and AIDS
      Boston, Massachusetts, United States