5′ upstream sequences of MyD88, an IL-6 primary response gene in M1 cells: detection of functional IRF-1 and Stat factors binding sites

Abstract

Transcription regulatory elements have been analyzed in upstream sequences of an Interleukin-6 (Il-6) primary response gene, MyD88. MyD88 2.3 kb mRNA is strongly and persistently induced in the course of myeloleukemic M1 cells differentiation with Il-6. MyD88 cDNA sequences were found in a region of 12 kb of mouse genomic DNA. Using Il-6 treated M1 cell RNAs, two transcription start sites have been localized, approximately 100 bp upstream from the 5' end of the cloned cDNA. We sequenced 1.4 kb of 5' genomic DNA including the first exon. In 5' of mRNA transcription start site, MyD88 nucleotidic sequence is 85% identical to 5' complementary sequences of the rat 3'-ketoacetyl CoA thiolase gene, over 1.2 kb. A DNA element conferring Il-6-inducible transcription to reporter genes, and localized 30 bp upstream of MyD88 first RNA start site, contains overlapping binding sites for cytokine activated transcription factors Stat and for the Interferon Regulatory Factor-1 and -2 (IRF-1 and IRF-2). In vitro binding assays showed that attachment of Stat factors to this element early in Il-6 treatment requires tyrosine kinase activation. IRF1, an activator of transcription, is also induced to bind to this sequence at later times. A model of persistent activation of MyD88 gene through these two types of factors is proposed.

Regulation of IKKβ by miR-199a affects NF-κB activity in ovarian cancer cells

“…Furthermore, IKKb overexpression in Type II cells led to the production of high levels of proinflammatory cytokines similar to that we saw in Type I cells ( Figure 4b). Interestingly, we also observed increased MyD88 expression in these transfectants (Figure 4a), which may be the result of NF-kB activation, as MyD88 is also a target of NF-kB (Harroch et al, 1995).…”

Endogenously Expressed nef Uncouples Cytokine and Chemokine Production from Membrane Phenotypic Maturation in Dendritic Cells

“…Subsequent studies examining LPS triggering through TLR4 (54) or dsRNA via TLR3 (55) further highlight how one stimulus-receptor interaction can signal distinct pathways within DCs, specifically MyD88 adaptor protein-dependent cytokine production vs MyD88-independent membrane phenotypic maturation. Interestingly, we observed nef-mediated Stat3 activation in immature DCs (35) that may favor MyD88-dependent cytokine secretion (56). The fact that nef, but not maturation stimuli, could rescue the defective replication of SIV delta nef in the immature DC-T cell mixtures (Fig.…”

Unmasking by soluble IL-6 receptor of IL-6 effect on metastatic melanoma: growth inhibition and differentiation of B16-F10.9 tumor cells

“…Other tumor cells with partial growth-inhibition by IL-6 become more inhibited when sIL-6R is added, for example human melanoma A375 (Oh et al, unpublished), human T47D breast cancer cells (Novick et al, 1992) and myeloleukemic M1 cells (Taga et al, 1989). Screening of primary and established tumor cells for the sIL-6R e ect may be of interest, even though resistance to IL-6 growth inhibition is not always overcome by sIL-6R (Silvani et al, 1995) and may be due to high IRF-2/IRF-1 ratios (Harroch et al, 1993(Harroch et al, , 1994a. Nevertheless, considering that about 50% of advanced melanomas produce autocrine IL-6 (Colombo et al, 1992;Lu and Kerbel, 1993;Armstrong et al, 1994), sIL-6R may enhance IL-6 e ects of many tumor cells which have lost surface IL-6R due to autocrine IL-6 (Silvani et al, 1995) either by retention of IL-6R inside the cell (Rose-John et al, 1993) or by decrease in IL-6R mRNA (Ganapathi et al, 1996).…”

“…Treated cells were washed twice in ice-cold phosphate bu ered saline (PBS) and pellets frozen in liquid N 2 . Nuclear extracts (Harroch et al, 1993(Harroch et al, , 1994a were used for EMSA performed with pIRE/IRF1, pIRE/a2M and C13 DNA probes as described before (Harroch et al, 1993(Harroch et al, , 1994a.…”