Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P05231 (interleukin-6)
 23,907 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have designed an expression vector for the secretion of human interleukin-6 (hIL-6) in which the mature protein is fused through a spacer peptide, containing a KEX-2 like protein processing signal, to the entire Aspergillus niger glucoamylase (glaA) gene. Transformation of Aspergillus nidulans with this vector results in fungal strains secreting equimolar amounts of the glucoamylase and IL-6 proteins. The KEX2-type processing signal, Lys-Arg, is recognized and cleaved efficiently by an enzyme present in A. nidulans resulting in the secretion of an authentic mature hIL-6 protein at levels of up to 5 mg/l.
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PMID:Efficient KEX2-like processing of a glucoamylase-interleukin-6 fusion protein by Aspergillus nidulans and secretion of mature interleukin-6. 136 12

IL-6-PE4E is a recombinant protein consisting of interleukin-6 (IL-6) fused to a mutant form of Pseudomonas exotoxin in which four basic amino acids are changed to glutamate (PE4E). The chimeric toxin has been previously shown to specifically kill malignant hepatic, prostatic, epidermoid, and myeloma cell lines in vitro. To explore the possible clinical utility of IL-6-PE4E, particularly as an agent for ex vivo purging of marrow for autologous bone marrow transplantation (ABMT), we tested malignant cells from patients with multiple myeloma for sensitivity to this chimeric toxin. Ficoll-purified bone marrow cells were incubated with and without IL-6-toxin for 2 to 3 days. Eight of the 15 myeloma patients had cells that were sensitive to IL-6-toxin as measured by a decrease in the level of protein synthesis. Cells from five patients were very sensitive to IL-6-PE4E, with 50% inhibition of protein synthesis (ID50) achieved at or below 6 ng/mL (7 x 10(-11) mol/L). Cells from three additional patients showed moderate sensitivity, with ID50s between 30 and 140 ng/mL. The remaining seven samples showed little or no sensitivity, with ID50s greater than or equal to 400 ng/mL. Normal bone marrow cells or normal BFU-E and CFU-GM were resistant to the IL-6-toxin even at 1,000 ng/mL. Neither IL-6, IL-2-PE4E, nor an enzymatically deficient mutant of IL-6-PE4E was cytotoxic toward the myeloma cells, indicating that the cytotoxic effect of IL-6-PE4E required the adenosine diphosphate-ribosylation function as well as the specific ligand. Our data suggest that IL-6-toxin could be effective in ex vivo marrow purging in selected multiple myeloma patients who are candidates for ABMT, and that this toxin should also be investigated further for in vivo therapy.
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PMID:Interleukin-6 fused to a mutant form of Pseudomonas exotoxin kills malignant cells from patients with multiple myeloma. 155 71

Rat T-kininogen (T-KG), a cysteine protease inhibitor, is an acute phase reactant which is induced to high levels in response to inflammation. Both hormones and cytokines participate in this regulation. To investigate the cis-acting elements responsible for the induction of gene expression, various 5'-fragments of the rat T-KG gene were fused to a chloramphenicol acetyltransferase marker gene. These constructs were transfected into a rat hepatoma cell line which was then treated with tumor necrosis factor or interleukin-6 or both cytokines. Expression of the chloramphenicol acetyltransferase gene was induced with interleukin-6 treatment, but suppressed by tumor necrosis factor. The 5'-region of the T-KG gene responsible for conferring both of these effects was localized between nucleotides -404 to -210 upstream of the transcription start site. Fragments containing this region were found to be effective in either orientation, and could also regulate a heterologous promoter.
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PMID:Differential regulation of rat T-kininogen by tumor necrosis factor and interleukin-6. 170

Interleukin-6 (IL-6) activates (2'-5') A synthetase (2'-5' AS) gene expression in differentiating myeloleukemic M1 cells. Antibodies to type I interferon (IFN) inhibit 2'-5' AS induction but not differentiation. Analysis of the mechanism of 2'-5' AS induction shows that it does not result from increased IFN formation, but from a synergism between IL-6 and endogenously secreted IFN. IL-6 can activate expression of a CAT construct fused to the interferon response sequence (IRS) of the 2'-5' AS gene. In extracts of IL-6-treated M1 cells, changes in protein binding to IRS DNA can be demonstrated. One of the effects of IL-6 on M1 cells is, therefore, to induce DNA binding factors, some of which act on the same enhancer sequence as IFNs, resulting in a synergistic gene activation. M1 variants resistant to differentiation by IL-6 have lost the ability to induce the 2'-5' AS gene.
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PMID:Interleukin-6 induces the (2'-5') oligoadenylate synthetase gene in M1 cells through an effect on the interferon-responsive enhancer. 188 86

The serum concentration of rat T1 kininogen increases 20-30-fold in response to acute inflammation, an induced hepatic synthesis regulated primarily at the transcriptional level. To analyze the cis-regulatory elements responsible for the induced transcription, we fused a 1.6-kilobase segment of the rat T1 kininogen promoter to a reporter gene, chloramphenicol acetyltransferase (CAT). The resultant chimeric DNA was transfected into cultured cells. In transient transfection assays, this 5'-flanking sequence was sufficient to confer cell-specific expression: CAT activity was readily detectable when the construct was transfected into liver-derived cells, but it was not detectable in nonliver cells. Furthermore, when liver cells (Hep3B) transfected with this construct were treated with conditioned medium prepared from activated mixed lymphocyte cultures or with recombinant interleukin-6 (IL-6), a 5-fold increase in CAT activity was detected. Addition of dexamethasone to the conditioned medium or to IL-6 showed synergistic effects and resulted in a 10-fold increase in CAT activity. In contrast, when IL-1 was used with IL-6, induction of CAT activity was inhibited. Deletion analyses revealed two regions important for tissue-specific and induced regulation of T1 kininogen: sequences proximal to base pair -73 conferred enhanced expression in liver-derived cells and a distal region that conferred responsiveness to conditioned medium, recombinant IL-6, and dexamethasone. This responsive element had properties of an inducible transcriptional enhancer, and it was functional in both liver and nonliver cells when placed immediately upstream of a thymidine kinase promoter.
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PMID:Interleukin-6 responsiveness and cell-specific expression of the rat kininogen gene. 199 68

The coding region of the human interleukin-6 (hIL6) gene was fused to the prepro secretion signal of the alpha-mating factor gene in several yeast host strains. It was found that the KEX-2 protease was unable to cleave the prepro-Lys-Arg-Pro-IL6 sequence, but that unspecific cleavage of the precursor protein had occurred. The prepro-Lys-Arg-Ala-Pro-IL6 sequence, however, was correctly recognized and cleaved by the KEX-2 protease, and IL6 was efficiently secreted into the culture medium. The N-terminal Ala-Pro peptide was removed during processing by wild-type yeast strains, but was retained in a ste13 mutant. IL6 as well as the aberrant proteins were not glycosylated. The transformed cells could secrete up to 30 micrograms/ml IL6. The protein was purified from the medium to homogeneity by ion-exchange chromatography and gel filtration, and had a specific activity of about 2 x 10(8) IU/mg in a proliferation assay.
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PMID:Production and purification of recombinant human interleukin-6 secreted by the yeast Saccharomyces cerevisiae. 204 Feb 82

To study the mechanism of induction of human C-reactive protein (CRP) gene expression, we have utilized an in vitro liver cell system to analyze the cis-acting DNA sequences located within the 5'-flanking region of human CRP gene. Stable transfection of human hepatoma cells, PLC/PRF/5, by a CRP gene construct containing the 1 kilobase pair of upstream sequence of the CRP gene demonstrated that this region contained the inducible element(s) which regulated human CRP gene transcription. Dissection of this region by 5', 3' and internal deletion constructs of upstream region of the CRP gene fused to a reporter gene, chloramphenicol acetyl transferase, indicated the presence of two inducible elements located proximal to the site of initiation of transcription, two constitutive enhancer-like elements located distal to the promoter, and a negative regulatory region located between the two inducible elements. We had previously shown that a protein factor from monocytes or HTLV1-infected T-cells, was responsible for CRP induction in hepatoma cells. We have found this factor to be synonymous with interleukin-6. By stable and transient transfection assays in hepatoma cells, recombinant interleukin-6 alone was sufficient to activate both inducible elements.
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PMID:cis-acting elements responsible for interleukin-6 inducible C-reactive protein gene expression. 215 96

Expression vectors for human interleukin-6 (hIL6) contain an expression cassette consisting of the Aspergillus niger glaA promoter and the Aspergillus nidulans argB terminator. The secretion signals were either those of glaA or that of the authentic hIL6 peptide. The constructs under study were introduced into A. nidulans and A. niger by means of cotransformation. No IL6 activity could be detected in the medium of a cotransformed A. niger strain, although transcripts corresponding with the IL6 cDNA were present. Evidence is presented that this apparent lack of IL6 expression is due to extracellular proteolytic activity. In the media of a cotransformed A. nidulans strain grown on starch, IL6 activity was detected by means of a bioassay. Up to 25 ng/ml of biologically active hIL6 could be secreted by A. nidulans transformed with the plasmid containing the mature hIL6-encoding gene fused to the glaA signal peptide nucleotide sequences. hIL6 of the expected 23-kDa size was also observed by Western-blot analysis of the medium. There was no evidence for glycosylation of the protein.
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PMID:Heterologous gene expression by filamentous fungi: secretion of human interleukin-6 by Aspergillus nidulans. 225 49

Two types of recombinant human IL-6 (rIL-6) were used for the development of specific monoclonal antibodies. The first was produced in E. coli and used for immunization, the second was produced in Chinese Hamster Ovary Cells (CHO) and used for screening. The complete translated sequence of the cDNA coding for human IL-6 was fused, in phase, to protein-A and the hybrid gene was fused to the strong lambda PR promoter. This protein was purified from bacterial extracts by chromatography on rabbit IgG-Sepharose columns. After six injections of the purified protein into mice, sera were tested for their binding titer in a solid phase radioimmunassay (sRIA) and for the specificity of binding by Western blots. In the sRIA, crude supernatants of CHO cells (harboring a plasmid containing the human IL-6 gene and expressing high levels of IL-6 but no protein-A or any bacterial antigen) were bound to a solid support, reacted with supernatants of the hybridomas and finally detected with [125I]-goat anti-mouse antibodies. Spleen cells derived from a mouse showing the highest binding titer were fused to mouse myeloma cells. The hybridomas were screened by the sRIA and several positive clones were isolated and characterized. One of the clones was found to neutralize the hybridoma growth factor activity of the rIL-6 from both sources. The same clone was also used for Western blots and for affinity purification of both natural and recombinant IL-6 (E. coli and CHO).
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PMID:Monoclonal antibodies for affinity purification of IL-6/IFN- beta 2 and for neutralization of HGF activity. 268 Sep 1

Transcription of the human haptoglobin (Hp) gene is induced by interleukin-6 (IL-6) in the human hepatoma cell line Hep3B. Cis-acting elements responsible for this response are localized within the first 186 bp of the 5'-flanking region. Site-specific mutants of the Hp promoter fused to the chloramphenicol acetyl transferase (CAT) gene were analysed by transient transfection into uninduced and IL-6-treated Hep3B cells. We identified three regions, A, B and C, defined by mutation, which are important for the IL-6 response. Band shift experiments using nuclear extracts from untreated or IL-6-treated cells revealed the presence of IL-6-inducible DNA binding activities when DNA fragments containing the A or the C sequences were used. Competition experiments showed that both sequences bind to the same nuclear factors. Polymers of oligonucleotides containing either the A or the C regions confer IL-6 responsiveness to a truncated SV40 promoter. The B region forms several complexes with specific DNA-binding proteins different from those which bind to the A and C region. The B region complexes are identical in nuclear extracts from IL-6-treated and untreated cells. While important for IL-6 induction in the context of the haptoglobin promoter, the B site does not confer IL-6 inducibility to the SV40 promoter. Our results indicate that the IL-6 response of the haptoglobin promoter is dependent on the presence of multiple, partly redundant, cis-acting elements.
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PMID:The human haptoglobin gene promoter: interleukin-6-responsive elements interact with a DNA-binding protein induced by interleukin-6. 278 45


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