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  • Title: Transcriptional regulation of the gilthead seabream (Sparus aurata) interleukin-6 gene promoter.
    Author: Castellana B, Marín-Juez R, Planas JV.
    Journal: Fish Shellfish Immunol; 2013 Jul; 35(1):71-8. PubMed ID: 23602849.
    Abstract:
    Interleukin-6 (IL-6) has been identified and characterized from several fish species and its mRNA expression is induced by pathogen-associated molecular patterns (PAMPs) and cytokines in immune cells and tissues. However, the transcriptional regulation of the IL-6 gene in fish is not well understood. In the present study, we have cloned and sequenced a 1028 bp 5'-flanking DNA region from the IL-6 gene in seabream (Sparus aurata). Sequence analysis of the seabream IL-6 promoter (sbIL-6P) evidenced the presence of a conserved TATA motif and conserved response elements for NF-κB, C/EBPβ (NF-IL6), AP-1 and GRE, similar to other vertebrate IL-6 promoters. Functional characterization of sbIL-6P was performed by cloning sbIL-6P into a luciferase expression vector and by transfecting it into L6 muscle cells, a mammalian cell line shown previously to express IL-6 in response to pro-inflammatory stimuli. We show here that the activity of sbIL-6P was significantly induced by pro-inflammatory cytokines such as tumor necrosis factor alpha (TNFα), IL-6 and IL-2, as well as by lipopolysaccharide (LPS), but significantly repressed by dexamethasone. In addition, the stimulatory effects of TNFα on sbIL-6P activity appeared to be mediated by the NF-κB, p38 MAPK and JNK signaling pathways. Deletion analyses of sbIL-6P suggested that activation of sbIL-6P by TNFα and IL-6 required the presence of binding motifs present in the proximal promoter (-171 to -84) whereas activation by IL-2 required binding motifs present in the distal promoter (-1024 to -864). The results from this study indicate, for the first time in fish, that pro-inflammatory cytokines, LPS and glucocorticoids can regulate the activity of the IL-6 gene at a transcriptional level and identify important regions in its response to cytokines.
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