Biomedical subjects
S Akira
Publications and source records attributed to S Akira.
The role of NF-kappa B and NF-IL6 transactivating factors in the synergistic activation of human serum amyloid A gene expression by interleukin-1 and interleukin-6.
To understand the mechanisms by which large increases in serum amyloid A (SAA) occur during the acute phase response, human hepatoma cells were transfected with SAA2 gene reporter plasmids and stimulated with combinations of cytokines. Although interleukin-1 (IL-1) and interleukin-6 (IL-6) stimulated transcription from this promoter individually, addition of both mediators produced a response between two and nine times greater than the expected additive response. This synergistic activation was dependent on the integrity of at least two cis-acting sequences in the SAA2 enhancer. The SAA2 NF-kappa B site was required functionally for the response to both IL-1 and IL-6 alone as well as for synergistic activation; however, IL-6 did not directly induce binding of nuclear proteins to the NF-kappa B sequence. A NF-IL6 site was required for full induction by IL-1 and IL-6, and also mediated strong transactivation by recombinant NF-IL6. Furthermore, transfected NF-IL6 synergized strongly with co-transfected NF-kappa B, particularly with RelA (p65). However synergy between IL-1 and IL-6 was only partly reduced by mutation of the NF-IL6 site, indicating further levels of interaction in addition to the NF-kappa B/NF-IL6 cooperativity.
Transcription factors NF-IL6 and NF-kappa B synergistically activate transcription of the inflammatory cytokines, interleukin 6 and interleukin 8.
Single binding sites for transcription factors NF-IL6 and NF-kappa B are present in the promoter of the interleukin (IL) 6 gene. Previous studies of internally deleted promoter mutants demonstrated that these two sites are important for the transcriptional regulation of this gene. In this report, we describe the synergistic activation of the IL-6 promoter by transcription factors NF-IL6 and NF-kappa B. Cotransfection of NF-IL6 with the NF-kappa B p65 subunit resulted in strong synergistic activation of an IL-6 promoter-reporter construct. Both the NF-IL6 and NF-kappa B binding sites in the IL-6 promoter were required for synergistic activation. Similar synergistic activation was observed in the IL-8 promoter, which also contains both NF-IL6 and NF-kappa B binding sites. Furthermore, we demonstrated that NF-IL6 and the NF-kappa B p65 subunit directly associated via the basic leucine-zipper domain of NF-IL6 and the Rel homology domain of p65. Since the promoters of many other genes involved in the inflammatory and acute-phase responses also contain binding sites for NF-IL6 and NF-kappa B, the cooperation between these two factors may have an important role in these responses. We also discuss the possible interplay between various viral gene products and these two factors in the process of viral infection and constitutive cytokine production.
Phosphorylation at threonine-235 by a ras-dependent mitogen-activated protein kinase cascade is essential for transcription factor NF-IL6.
NF-IL6, a member of the basic leucine zipper (bZIP) family transcription factors, is involved in expression of inducible genes involved in immune and inflammatory responses. We observed that coexpression of oncogenic p21ras stimulated the transactivating activity of NF-IL6 and induced phosphorylation of Thr-235 located just N-terminal to the DNA binding domain of NF-IL6. Recently, mitogen-activated protein (MAP) kinases have been shown to be implicated in the cellular response to activated ras. Purified MAP kinases specifically phosphorylated Thr-235 of NF-IL6 in vitro. Mutation of Thr-235 abolished the ras-dependent activation of NF-IL6. From these results, we conclude that NF-IL6 is regulated through phosphorylation by MAP kinases in response to activated ras.
Intussusception caused by a carcinoma of the cecum during pregnancy: report of a case and review of the literature.
A case of intussusception due to a carcinoma of the cecum during pregnancy is reported. A 27-year-old pregnant female was admitted to Shimodate Municipal Hospital because of abdominal pain, nausea and vomiting. Her abdomen was distended, and a relatively hard mass was palpable in the right hypochondrium. Following a diagnosis of intussusception by ultrasonography, a laparotomy was performed. The lesion causing the intussusception was found to be a carcinoma of the cecum, and thus a right hemicolectomy with lymph node dissection was carried out. Histological examination revealed that the tumor was a well-differentiated adenocarcinoma which had invaded the muscularis propria but was superficial to the subserosa. None of the lymph nodes were cancerous. The incidence of colonic cancer above the peritoneal reflection during pregnancy is very low. Only 24 cases have been previously reported; our patient is only the 25th case, as well as being the first case demonstrating Dukes' A. Due to the intussusception, ultrasonography was effective for diagnosis and the patient was able to undergo a curative operation at an earlier stage than other patients.
Pregnancy can be established in superovulated adult rats treated with progesterone and an aromatase inhibitor.
An adult superovulated rat model has been developed and is characterized by high ovulation rates, early morphological degeneration of embryos, complete embryo loss within 48 hours of conception and elevated peripheral estradiol(E2)/progesterone(P4) ratios. In this study, three trials were conducted using the superovulated adult rat model. First, control naturally cycling rats were compared with superovulated rats supplemented with 1 mg P4 on days 0-3 of pregnancy. A sperm positive vaginal smear is designated as day 0 of pregnancy. The P4 treated rats demonstrated improved embryo retrieval on day 1 of pregnancy, continued embryo recovery with a decrease in normal morphologic characteristics of integrity on day 2, with nearly total embryo loss by day 3. On each day, P4 levels were elevated 2-3 times over control. The second trial compared 3 groups of rats, 1) naturally cycling, 2) superovulated unsupplemented and 3) superovulated rats supplemented with 1 mg P4/rat/day and the aromatase inhibitor, 4-hydroxyandrostenedione (4-OHA), 12.5 mg/rat/day. The superovulated unsupplemented rats had no embryo recovery after day 2 of pregnancy, while the P4 and 4-OHA treated rats showed a variable ability to maintain normally developing embryos through day 4 of pregnancy. E2 levels were elevated in both superovulated groups on days 1-4 of pregnancy as were P4 levels on days 2-4. The E2/P4 ratio was significantly lowered only on day 1 of pregnancy in the P4 and 4-OHA treated group. The third trial demonstrated implantation in 50% of the superovulated rats supplemented with P4 and 4-OHA. In conclusion, implantation in the superovulated adult rats can occur with P4 and 4-OHA supplementation, however, this biologic phenomenon could not be explained by obvious changes in peripheral E2 and P4 levels.
Acceleration of embryo transport in superovulated adult rats.
The etiology of reduced fertility in rodents and humans after exogenous gonadotropin-induced superovulation is unknown. This study examines implantation failure in adult rats induced to superovulate by gonadotropin treatment. After multiple injections of pregnant mare serum gonadotropin (PMSG) followed by human chorionic gonadotropin (hCG), superovulation in adult rats was confirmed on day-1 of pregnancy, however, on day-13, no implantation sites were observed. Daily flushing of oviducts and uteri revealed that the number of embryos recovered from superovulated rats on day-2 was only one-third of that retrieved on day-1. The numbers of embryos retrieved on these days from the oviducts of control rats, however, were almost equivalent. On day-3, no embryos were retrieved from either the oviducts or uteri of superovulated rats, while all embryos from control rats were found in the oviducts. Most embryos recovered from superovulated rats showed normal development. Compared to levels in control rats, serum estradiol (E2) in superovulated rats increased significantly on days-2 and -3 of pregnancy, whereas, serum progesterone (P) levels remained unchanged. Thus, E2/P ratio increased on days-2 and -3 of pregnancy in superovulated adult rats coinciding with the timing of embryo transport acceleration. Our results suggest that superovulated oocytes can be fertilized and reach an early stage of development within the oviduct. Implantation failure in superovulated rats may be due to the accelerated embryo transport resulting from elevated E2/P ratio.
Interleukin-6 in biology and medicine.
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NF-IL6 represses early gene expression of human papillomavirus type 16 through binding to the noncoding region.
The expression of human papillomavirus type 16 (HPV16) early genes, including E6 and E7 transforming genes, is regulated by several cellular factors binding to the noncoding region (NCR), such as the glucocorticoid receptor, NF-I, and AP1, all of which are positive regulators. We demonstrated that the nuclear factor for interleukin 6 expression (NF-IL6) specifically binds to the HPV16 NCR ranging from nucleotides 7007 to 7766 and represses the early gene expression of HPV16. The responsive element in HPV16 NCR was determined within the region ranging from nucleotides 7454 to 7766. In this region, many binding sites for other cellular transactivators, such as NF-I and AP1, have been detected. Interestingly, three of seven binding sites for NF-I and two of two binding sites for AP1 in this region overlap with the putative NF-IL6 binding sites identified by computer analysis. Competition experiments with the oligonucleotides containing such NF-I or AP1 sites indicated that NF-IL6 certainly binds to them. Furthermore, in a chloramphenicol acetyltransferase assay using mutant NF-IL6 expression vectors, the DNA binding domain of NF-IL6 was shown to be necessary for repression, whereas the functional domain was not. These findings indicate that repression may be caused by competition with other transcriptional activators, such as NF-I and AP1. Thus, NF-IL6 may play a significant role in the regulation of viral transcription as a part of the host's resistance to viral infection.
A nuclear factor for interleukin-6 expression (NF-IL6) and the glucocorticoid receptor synergistically activate transcription of the rat alpha 1-acid glycoprotein gene via direct protein-protein interaction.
The acute-phase reaction is accompanied by an increase in a variety of serum proteins, named acute-phase proteins. The synthesis of these proteins is synergistically controlled by glucocorticoids and inflammatory cytokines such as interleukin-1 (IL-1), IL-6, and tumor necrosis factor alpha. Recently, we have cloned nuclear factor-IL-6 (NF-IL6), a transcription factor that activates the IL-6 gene, and have demonstrated its involvement in the expression of acute-phase-protein genes. We report here an analysis of the molecular mechanisms by which inflammatory cytokines and glucocorticoid act synergistically to activate expression of the rat alpha 1-acid glycoprotein (AGP) gene. We found that NF-IL6 and ligand-activated rat glucocorticoid receptor acted synergistically to transactivate the AGP gene and that maximal transcriptional activation of the AGP gene required expression of both intact NF-IL6 and rat glucocorticoid receptor. Surprisingly, however, transcriptional synergism was still observed even when one of the two factors lacked either its DNA-binding or transcriptional-activation function. We present evidence for a direct protein-protein interaction between these two distinct transcription factors and propose that this may be responsible for the synergistic activation of the rat AGP gene.
Interleukin-6 and its receptor: a paradigm for cytokines.
Many cytokines and cytokine receptors involved in the regulation of hematopoiesis, immune responses, and inflammation have been identified and characterized at the molecular level. Several characteristic features of cytokines, such as pleiotropy and redundancy, are now more clearly understood on the basis of their molecular structures. Accumulating evidence has demonstrated an intimate link between cytokines and various diseases such as allergy, autoimmune diseases, and cancer. The pathogenesis of these diseases and therapies to treat them will be discussed based on insights derived from cytokine research.
A member of the C/EBP family, NF-IL6 beta, forms a heterodimer and transcriptionally synergizes with NF-IL6.
Using a DNA probe from the DNA-binding portion of the NF-IL6 gene and an antibody against the DNA-binding domain of NF-IL6, we isolated a gene homologous to NF-IL6 in the DNA-binding and leucine zipper domains. This intronless gene, termed NF-IL6 beta encodes a 269-amino acid protein with a potential leucine zipper structure, and the gene product can bind to the CCAAT homology as well as the viral enhancer core sequence, as in the cases of NF-IL6 and C/EBP. This gene is expressed at an undetectable or a minor level in normal tissues but is induced by lipopolysaccharide or inflammatory cytokines, as in the case of NF-IL6. NF-IL6 beta easily forms a heterodimer with NF-IL6 in vitro and the heterodimeric complex binds to the same DNA sequence as the respective homodimers. When examined by transient luciferase assays, NF-IL6 beta is consistently a stronger transactivator than NF-IL6. Furthermore, NF-IL6 beta shows a synergistic transcriptional effect with NF-IL6. These data suggest that NF-IL6 beta is an important transcriptional activator in addition to NF-IL6 in regulation of the genes involved in the immune and inflammatory responses.
Macrophage differentiation-specific expression of NF-IL6, a transcription factor for interleukin-6.
NF-IL6 was originally identified as a DNA binding protein regulating interleukin-1 (IL-1)-stimulated IL-6 expression. Direct cloning of NF-IL6 showed its homology with C/EBP, a hepatocyte- and adipocyte-specific transcription factor. This study showed that the expression of NF-IL6 messenger RNA (mRNA) increased markedly during the differentiation to a (mRNA) increased markedly during the differentiation to a macrophage lineage in mouse myeloid leukemia cells M1, human histiocytic leukemia cells U937, promyelocytic leukemia cells HL-60, and human peripheral monocytes. Particularly in HL-60 cells that undergo granulocyte or macrophage differentiation depending on inducers, NF-IL6 mRNA was specifically upregulated during macrophage differentiation but not granulocyte differentiation. It was also shown that the functional NF-IL6 protein increased during the differentiation of U937 cells. Furthermore, recombinant NF-IL6 was found to bind to the regulatory regions of the IL-1, tumor necrosis factor, granulocyte colony-stimulating factor, and lysozyme genes, which are expressed in mature macrophages. These results suggest that NF-IL6 may possibly be involved as an important transcription factor in the process of activation and/or differentiation of macrophages.
Regulation of expression of the interleukin 6 gene: structure and function of the transcription factor NF-IL6.
The interleukin 6 (IL-6) promoter is rapidly and transiently activated by other cytokines, including IL-1 and tumour necrosis factor (TNF), as well as by phorbol esters and cyclic AMP agonists. Studies using promoter mutants suggested that an IL-1-responsive element mapped within the -180 to -123 region of the IL-6 promoter. A nuclear factor (NF-IL6) that recognized a unique sequence containing an inverted repeat, ACATTGCACAATCT, was identified within the region. Direct cloning of the human NF-IL6 revealed its similarity to C/EBP, a liver- and adipose tissue-specific transcription factor. C/EBP and NF-IL6 recognize the same nucleotide sequence, but exhibit distinct patterns of expression. NF-IL6 is expressed at a low level in normal tissues, but is rapidly and drastically induced by bacterial lipopolysaccharide (LPS) or inflammatory cytokines such as IL-1, TNF and IL-6. Recently, NF-IL6 has been shown to be identical to IL-6DBP, the DNA-binding protein which is responsible for IL-6-mediated induction of several acute-phase proteins. Evidence that NF-IL6 DNA-binding activity is increased after IL-6 stimulation without increased NF-IL6 protein synthesis demonstrates the importance of post-translational modification. There are some results indicating that phosphorylation is involved in transcriptional and binding activities of NF-IL6. Taken together, these findings indicate that NF-IL6 may be an important transcription factor on the signal transduction pathways of IL-1 and IL-6.
Stimulatory effect of thymic factor(s) on steroidogenesis in cultured rat granulosa cells.
Thymic cells from immature female rats were isolated and used for production of thymic cell culture conditioned medium (TCM). Granulosa cells were obtained from immature diethylstilbestrol (DES)-treated rats. TCM stimulated basal progesterone and estradiol secretion from the granulosa cells in a dose and time dependent manner. Maximal stimulation of progesterone production occurred at 48 hours of incubation, during which period TCM caused approximately 5 times more progesterone secretion than heart cell conditioned medium (HCM) or mock extract (ME). The maximum progesterone secretion by granulosa cells occurred when they were exposed to 48% TCM causing 7 times more progesterone secretion than controls. Under the same maximum stimulatory conditions, however, TCM only approximately doubled estradiol secretion compared to concentrations secreted in the presence of HCM or ME. Thus, the effect of TCM on progesterone secretion was more prominent than its effect on estradiol secretion. The stimulatory action of TCM was not mimicked by HCM, thymosin-alpha 1 or thymulin. Furthermore, the stimulatory action of TCM on steroidogenesis did not appear to be mediated by the cAMP system. The stimulatory factor(s) in TCM were heat, acid and acetone labile, but could not be sedimented by activated charcoal. Thus, the present study demonstrates that the secretory product(s) of thymic epithelial cells can stimulate steroidogenesis in cultured rat granulosa cells. Our data imply that thymic factor(s) may have a direct effect on ovarian function.
IL-6 receptor and mechanism of signal transduction.
Interleukin-6 (IL-6) is a multifunctional cytokine which acts on a wide variety of cells, exerting growth promotion, growth inhibition, or specific gene expression including cellular differentiation. The IL-6 receptor system consists of two membrane proteins, a ligand-binding chain (IL-6R) and a non-ligand-binding signal transducer, gp130, both of which belong to the cytokine receptor family. Binding of IL-6 to IL-6R triggers the association of IL-6R and gp130, and gp130 in turn transduces the signal. Despite its lack of IL-6 binding property, gp130 is involved in the formation of high-affinity IL-6 binding sites. This two-chain IL-6 receptor system can be applied to some other cytokine receptors, such as IL-3R, IL-5R and GM-CSFR which share a second signal-transducing component. A nuclear factor for controlling IL-6 gene expression (NF-IL6) is a leucine zipper-containing transcription factor and is homologous to C/EBP, a liver nuclear factor. NF-IL6 is also involved in the transcriptional regulation of various acute phase protein genes IL-6-triggered association of IL-6R and gp130 on hepatocytes, through intermediate steps including serine-phosphorylation of pre-existing NF-IL6 protein, leads to binding of NF-IL6 to IL-6-responsive elements and activation of acute-phase protein genes.
NF-IL6 and gene regulation.
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Mechanisms of soluble mediators.
A two chain model, composed of ligand-binding and signal transducing subunits, first identified in the interleukin-6 receptor system, has been revealed in several other cytokine systems. Common signal transducers shared by several receptor systems may explain a functional redundancy characteristic of cytokines. The signaling pathway from the receptor to gene activation is now becoming clearer due to the successful molecular cloning of several transcription factors.