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S C Sun

Publications and source records attributed to S C Sun.

At least 55 records · Page 3Linked to original sources
PubMed1995

Dependence and reversal of nitric oxide production on NF-kappa B in silica and lipopolysaccharide-induced macrophages.

In this report the differential regulation of NF-kappa B and nitric oxide (NO) was investigated in the mouse macrophage cell line RAW 264.7 following exposure to a mineral dust (silica) and/or an endotoxin (Lipopolysaccharide, LPS). The results indicated that silica and LPS can significantly induce the activation of NF-kappa B as well as elicit enhanced production of NO in RAW 264.7 cells as part of an early inflammatory response mechanism. A 24-hour time-course study showed that NO release from these cells continued to increase following the initial stimulus by LPS or silica. In contrast, activation of NF-kappa B was maximal at 6 hours and then showed a steady decline to 24 hours. The production of NO was suppressed by protease inhibitor and antioxidant, both of which block the activation of NF-kappa B. Surprisingly, the use of an NO synthase inhibitor resulted in an enhancement of NF-kappa B activation. These findings suggest that NO produced in macrophage cells in response to an inflammatory stimulus like silica or LPS my be linked to a negative feedback role on the activation of NF-kappa B.

Acetylcysteine↗
PubMed1995

Essential role of NF-kappa B activation in silica-induced inflammatory mediator production in macrophages.

In this report, we demonstrate that NF-kappa B, a ubiquitous transcription factor, plays an essential role in silica-induced inflammatory mediator production in the mouse macrophage cell line RAW 264.7. Compared to the effect of lipopolysaccharide (LPS), silica mediated a stronger activation of NF-kappa B p50/p50 homodimer at early phase of poststimulation. Furthermore, activation of NF-kappa B by silica and LPS appears to be mediated by different signal transduction pathways. Both silica and LPS increased mRNA expression in these cells for cyclooxygenase II, inducible nitric oxide synthase, tumor necrosis factor-alpha and interleukin-1 alpha. This expression was attenuated along with the inhibition of NF-kappa B activation.

Animals↗
PubMed1995

Activation of NF-kappa B/Rel by Tax involves degradation of I kappa B alpha and is blocked by a proteasome inhibitor.

The tax gene product of the human T-cell leukemia virus type I (HTLV-I) induces the nuclear expression and biological function of the NF-kappa B/Rel family of host transcription factors although the underlying mechanism remains unclear. In the present study, we demonstrate that Tax-mediated activation of NF-kappa B/Rel can be inhibited by a proteasome inhibitor, suggesting the involvement of proteolytic reactions in this Tax-specific activation pathway. Transient transfection and reporter gene assays have revealed that Tax overrides the inhibitory function of I kappa B alpha in both F9 embryonal cells and Jurkat T cells. Moreover, Tax-mediated inactivation of I kappa B alpha requires a 16 amino acid sequence element located at the N-terminal region (amino acid 21-36) of I kappa B alpha, which is also required for tumor necrosis factor alpha-induced degradation of this inhibitory protein. We further demonstrate that the proteasome inhibitor also blocks the degradation of I kappa B alpha observed in HTLV-I-infected T cells. Interestingly, inhibition of I kappa B alpha degradation in these cells led to the accumulation of a phosphorylated form of I kappa B alpha. Together, these studies suggest that Tax activation of NF-kappa B/Rel may involve induction of phosphorylation and subsequent proteasome-mediated degradation of the inhibitor I kappa B alpha.

Amino Acid Sequence↗
PubMed1995

Activation of NF-kappa B by phosphatase inhibitors involves the phosphorylation of I kappa B alpha at phosphatase 2A-sensitive sites.

Activation of NF-kappa B by various cellular stimuli involves the phosphorylation and subsequent degradation of its inhibitor, I kappa B alpha, although the underlying mechanism remains unclear. In the present study, the role of serine/threonine phosphatases in the regulation of I kappa B alpha phosphorylation was investigated. Our studies demonstrate that incubation of human T cells with low concentrations (approximately 1-5 nM) of calyculin A or okadaic acid, potent inhibitors of protein phosphatase type 1 (PP-1) and type 2A (PP-2A), induces the phosphorylation of I kappa B alpha even in the absence of any cellular stimulus. This action of the phosphatase inhibitors, which is associated with the activation of the RelA.p50 NF-kappa B heterodimer, is not affected by agents that block the induction of I kappa B alpha phosphorylation by tumor necrosis factor alpha (TNF-alpha). Furthermore, the phosphorylated I kappa B alpha from calyculin A-treated cells, but not that from TNF-alpha-stimulated cells, is sensitive to PP-2A in vitro, suggesting the existence of fundamental differences in the phosphorylation of I kappa B alpha induced by the two different NF-kappa B inducers. However, induction of I kappa B alpha phosphorylation by both TNF-alpha and the phosphatase inhibitors is associated with the subsequent degradation of I kappa B alpha. We further demonstrate that TNF-alpha- and calyculin A-induced I kappa B alpha degradation exhibits similar but not identical sensitivities to a proteasome inhibitor. Together, these results suggest that phosphorylation of I kappa B alpha, mediated through both the TNF-alpha-inducible and the PP-2A-opposing kinases, may serve to target I kappa B alpha for proteasome-mediated degradation.

Cysteine Endopeptidases↗
PubMed1995

Transcription of immune genes in the giant silkmoth, Hyalophora cecropia, is augmented by H2O2 and diminished by thiol reagents.

Insects have an effective humoral immune system, residing in immune proteins that are synthesized largely in the fatbody. kappa B-like motifs upstream of the immune protein genes bind the Cecropia immunoresponsive factor (CIF), and confer high levels of inducible expression. We have extended our studies and sought evidence that oxygen-derived active species might modulate the expression of the immune protein genes and the activation of CIF after the administration of different inducers. Mitogens, like arachidonic acid, phorbol esters, phytohemagglutinin, bacteria, and components of microbial cell envelopes stimulate expression of the attacin genes, both in vivo and in vitro. A general stimulant of oxidative stress, H2O2, stimulated expression of these genes and the weak immune response to wounding was greatly augmented by administration of H2O2. These responses were largely or entirely inhibited by dithiothreitol and by N-acetylcysteine. Nonspecific responses were excluded since immune genes failed to respond to albumin or starch, and the expression of a non-immunoresponsive gene was not affected. Electrophoretic mobility-shift assays showed that H2O2 and bacteria, when administered in vitro, could activate CIF in fatbody cells and that dithiothreitol and N-acetylcysteine prevented this process. Our data suggest that the induction of the immune protein genes is mediated through the activation of CIF, contingent upon thiol oxidation induced by oxidative stress.

Acetylcysteine↗
PubMed1995

Structure and expression of Hemolin, an insect member of the immunoglobulin gene superfamily.

Hemolin is an insect protein which belongs to the immunoglobulin superfamily and is strongly induced upon bacterial infection. It has been isolated from two moths, Hyalophora cecropia and Manduca sexta. We have isolated and sequenced a genomic clone for hemolin in H. cecropia, in order to resolve its organization and as a basis for investigating hemolin gene regulation. According to Southern-blot analysis, hemolin is encoded by a single gene, Hemolin. It contains six exons ranging over 32-603 bp. The introns are positioned both within and between the immunoglobulin-like domains, a feature typical for cell-adhesion molecules belonging to the immunoglobulin superfamily. By an RNase protection assay, we show that the Hemolin transcript is strongly induced not only by bacteria, but also by lipopolysaccharide and phorbol 12-myristate 13-acetate. Analysis of the upstream region and introns revealed potential binding sites for the Cecropia immunoresponsive factor (CIF), which recognizes the kappa B-like consensus GGGRA YYYYY.

Amino Acid Sequence↗
PubMed1994

Autoregulation of the NF-kappa B transactivator RelA (p65) by multiple cytoplasmic inhibitors containing ankyrin motifs.

RelA (p65) functions as the critical transactivating component of the heterodimeric p50-p65 NF-kappa B complex and contains a high-affinity binding site for its cytoplasmic inhibitor, I kappa B alpha. After cellular activation, I kappa B alpha is rapidly degraded in concert with the induced nuclear translocation of NF-kappa B. The present study demonstrates that tumor necrosis factor alpha-induced degradation of I kappa B alpha in human T cells is preceded by its rapid phosphorylation in vivo. However, these effects on I kappa B alpha result in nuclear mobilization of only a fraction of the entire cytoplasmic pool of RelA. Subsequent studies have revealed that (i) cytoplasmic RelA is stably associated not only with I kappa B alpha but also with other ankyrin motif-rich proteins including the products of the NF-kappa B2 (p100) and NF-kappa B1 (p105) genes; (ii) in contrast to RelA-I kappa B alpha, RelA-p100 cytoplasmic complexes are not dissociated following tumor necrosis factor alpha activation; (iii) p100 functions as a potent inhibitor of RelA-mediated transcription in vivo; (iv) the interaction of RelA and p100 involves the conserved Rel homology domain of both proteins but not the nuclear localization signal of RelA, which is required for I kappa B alpha binding; (v) p100 inhibition of RelA function requires the C-terminal ankyrin motif domain, which mediates cytoplasmic retention of RelA; and (vi) as observed with I kappa B alpha, nuclear RelA stimulates p100 mRNA and protein expression. These findings thus reveal the presence of a second inducible autoregulated inhibitory pathway that helps ensure the rapid but transient action of nuclear NF-kappa B.

Ankyrins↗
PubMed1994

Human T-cell leukemia virus type I Tax activation of NF-kappa B/Rel involves phosphorylation and degradation of I kappa B alpha and RelA (p65)-mediated induction of the c-rel gene.

The tax gene product of human T-cell leukemia virus type I (HTLV-I) is a potent transcriptional activator that both stimulates viral gene expression and activates an array of cellular genes involved in T-cell growth. Tax acts indirectly by inducing or modifying the action of various host transcription factors, including members of the NF-kappa B/Rel family of enhancer-binding proteins. In resting T cells, many of these NF-kappa B/Rel factors are sequestered in the cytoplasm by various ankyrin-rich inhibitory proteins, including I kappa B alpha. HTLV-I Tax expression leads to the constitutive nuclear expression of biologically active NF-kappa B and c-Rel complexes; however, the biochemical mechanism(s) underlying this response remains poorly understood. In this study, we demonstrate that Tax-stimulated nuclear expression of NF-kappa B in both HTLV-I-infected and Tax-transfected human T cells is associated with the phosphorylation and rapid proteolytic degradation of I kappa B alpha. In contrast to prior in vitro studies, at least a fraction of the phosphorylated form of I kappa B alpha remains physically associated with the NF-kappa B complex in vivo but is subject to rapid degradation, thereby promoting the nuclear translocation of the active NF-kappa B complex. We further demonstrate that Tax induction of nuclear c-Rel expression is activated by the RelA (p65) subunit of NF-kappa B, which activates transcription of the c-rel gene through an intrinsic kappa B enhancer element. In normal cells, the subsequent accumulation of nuclear c-Rel acts to inhibit its own continued production, indicating the presence of an autoregulatory loop. However, the pathologic action HTLV-I Tax leads to the deregulated and sustained nuclear expression of both NF-kappa B and c-Rel, a response that may contribute to HTLV-I-induced T-cell transformation.

Base Sequence↗
PubMed1994

Human T-cell leukemia virus type I Tax associates with and is negatively regulated by the NF-kappa B2 p100 gene product: implications for viral latency.

Human T-cell leukemia virus type I (HTLV-I) is the etiologic agent of the adult T-cell leukemia, an aggressive and often fatal malignancy of activated human CD4 T cells. HTLV-I encodes an essential 40-kDa protein termed Tax that not only transactivates the long terminal repeat of this retrovirus but also induces an array of cellular genes. Tax-mediated transformation of T cells likely involves the deregulated expression of various cellular genes that normally regulate lymphocyte growth produced by altered activity of various endogenous host transcription factors. In particular, Tax is capable of modulating the expression or activity of various host transcription factors, including members of the NF-kappa B/Rel and CREB/ATF families, as well as the cellular factors HEB-1 and p67SRF. An additional distinguishing characteristic of HTLV-I infection is the profound state of viral latency that is present in circulating primary leukemic T cells. In this study, we demonstrate that HTLV-I Tax can physically associate with p100, the product of the Rel-related NF-kappa B2 gene, both in transfected cells and in HTLV-I-infected leukemic T-cell lines. Furthermore, the physical interaction of Tax with p100 leads to the inhibition of Tax-induced activation of the HTLV-I and human immunodeficiency virus type 1 long terminal repeats, reflecting p100-mediated cytoplasmic sequestration of the normally nuclearly expressed Tax protein. In contrast, a mutant of Tax that selectively fails to activate nuclear NF-kappa B expression does not associate with p100. Together, these results suggest that the cytoplasmic interplay of Tax and p100 may play an important role in the initiation and maintenance of HTLV-1 latency observed in adult T-cell leukemia.

Amino Acid Sequence↗
PubMed1993

kappa B-like motifs regulate the induction of immune genes in Drosophila.

The mammalian transcription factor NF-kappa B regulates a number of genes involved in immune and acute phase responses, by interacting with a nucleotide sequence element, the kappa B-motif. In this work we demonstrate the participation of similar motifs in the immune response of insects as well: kappa B-like motifs have a regulatory role in the synthesis of cecropins, a set of anti-bacterial peptides, triggered by the presence of bacterial cell wall components in the insect blood. We show that the upstream region of the Cecropin gene CecA1 contains elements responsible for inducible and tissue-specific expression. Furthermore, a trimer of kappa B-like motif confers high levels of inducible expression from the reporter gene, after transfection in a Drosophila blood cell line. As in the moth Hyalophora cecropia, stimulation with bacterial lipopolysaccharide induces a nuclear factor that specifically binds to the kappa B-like motif. Our data suggest a functional and evolutionary relationship between these insect immune response factors and the mammalian NF-kappa B.

Animals↗
PubMed1993

NF-kappa B controls expression of inhibitor I kappa B alpha: evidence for an inducible autoregulatory pathway.

The eukaryotic transcription factor nuclear factor-kappa B (NF-kappa B) participates in many parts of the genetic program mediating T lymphocyte activation and growth. Nuclear expression of NF-kappa B occurs after its induced dissociation from its cytoplasmic inhibitor I kappa B alpha. Phorbol ester and tumor necrosis factor-alpha induction of nuclear NF-kappa B is associated with both the degradation of performed I kappa B alpha and the activation of I kappa B alpha gene expression. Transfection studies indicate that the I kappa B alpha gene is specifically induced by the 65-kilodalton transactivating subunit of NF-kappa B. Association of the newly synthesized I kappa B alpha with p65 restores intracellular inhibition of NF-kappa B DNA binding activity and prolongs the survival of this labile inhibitor. Together, these results show that NF-kappa B controls the expression of I kappa B alpha by means of an inducible autoregulatory pathway.

CD4-Positive T-Lymphocytes↗
PubMed1993

The c-rel protooncogene product represses NF-kappa B p65-mediated transcriptional activation of the long terminal repeat of type 1 human immunodeficiency virus.

The long terminal repeat (LTR) of the type 1 human immunodeficiency virus (HIV-1) and the 5' regulatory region of the gene encoding the interleukin 2 receptor alpha subunit (IL-2R alpha) share functional kappa B enhancer elements involved in the regulation of these inducible transcription units during T-cell activation. These kappa B enhancer elements are recognized by a structurally related family of interactive proteins that includes p50, p65, and the product of the c-rel protooncogene (c-Rel). Recent biochemical studies have shown that p65 and p50 form the prototypical NF-kappa B complex, which is rapidly translocated from the cytoplasm to the nucleus during T-cell activation. This intracellular signaling complex potently stimulates kappa B-directed transcription from either the HIV-1 LTR or the IL-2R alpha promoter via the strong transactivation domain present in p65. We now demonstrate that nuclear expression of human c-Rel, which is induced by either phorbol ester or tumor necrosis factor alpha with delayed kinetics relative to p65, markedly represses p65-mediated activation of these transcription units. These inhibitory effects of c-Rel correlate with its DNA-binding activity but not with its ability to heterodimerize with p50, suggesting that c-Rel inhibition involves competition with p50/p65 for occupancy of the kappa B enhancer element. Together, these findings suggest that one function of c-Rel is as a physiologic repressor of the HIV-1 LTR and IL-2R alpha promoters, serving to efficiently counter the strong transcriptional activating effects of p65.

Base Sequence↗
PubMed1993

An ultrastructural observation of macrophages in rabbits' radius callus.

The ultrastructure of macrophages in rabbits' radius callus at the 14th and 31st day after fracture had been observed in the study. It was found that macrophages mostly appeared in the early stage callus where there were inflammation, clot, and bone chips. Commonly seen macrophages were active phagocytes with single nucleus, but they could evolve to form multinuclear macrophages or fuse into foreign body giant cells. We are therefore led to conclude that the functions performed by macrophages in healing callus are to phagocytize and digest damaged cellular fragments, small bone chips, clot and foreign body; to disrupt larger dead bone matrix extracellularly and to resorb them by microvilli-like processes on the surface of foreign body giant cells.

Animals↗
PubMed1993

A novel NF-kappa B complex containing p65 homodimers: implications for transcriptional control at the level of subunit dimerization.

The predominant inducible form of the NF-kappa B transcription factor is a heteromeric complex containing two Rel-related DNA-binding subunits, termed p65 and p50. Prior transfection studies have shown that when these p65 and p50 subunits are expressed independently as stable homodimers, p65 stimulates kappa B-directed transcription, whereas p50 functions as a kappa B-specific repressor. While authentic p50 homodimers (previously termed KBF1) have been detected in nuclear extracts from nontransfected cells, experimental evidence supporting the existence of p65 homodimers in vivo was lacking. We now provide direct biochemical evidence for the presence of an endogenous pool of inducible p65 homodimers in intact human T cells. As with the prototypical NF-kappa B p50-p65 heterodimer, this novel p65 homodimeric form of NF-kappa B is functionally sequestered in the cytoplasm but rapidly appears in the nuclear compartment following cellular stimulation. Site-directed mutagenesis studies indicate that the homodimerization function of p65 is dependent upon the presence of cysteine 216 and a conserved recognition motif for protein kinase A (RRPS; amino acids 273 to 276), both of which reside within a 91-amino-acid segment of the Rel homology domain that mediates self-association. In contrast, mutations at these two sites do not affect heterodimerization of p65 with p50 or its functional interaction with I kappa B alpha. These later findings indicate that neither homo- nor heterodimer formation is an absolute prerequisite for I kappa B alpha recognition of p65. Taken together with prior in vivo transcription studies, these results suggest that the biological activities of p65 and p50 homodimers are independently regulated, thereby providing an integrated and flexible control mechanism for the rapid activation and repression of NF-kappa B/Rel-directed gene expression.

Amino Acid Sequence↗