title: Transcriptional regulation of the gilthead seabream (Sparus aurata) interleukin-6 gene promoter.
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.
authors.affiliation: Departament de Fisiologia i Immunologia, Facultat de Biologia, Universitat de Barcelona and Institut de Biomedicina de la Universitat de Barcelona (IBUB), 08028 Barcelona, Spain.
authors.forename: Bàrbara
authors.initials: B
authors.lastname: Castellana
authors.forename: Rubén
authors.initials: R
authors.lastname: Marín-Juez
authors.forename: Josep V
authors.initials: JV
authors.lastname: Planas
mesh_terms: D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D029941:Fish Proteins / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D005938:Glucocorticoids / Q000235:genetics / Q000378:metabolism; D015850:Interleukin-6 / Q000235:genetics* / Q000378:metabolism; D008070:Lipopolysaccharides / Q000502:physiology; D008156:Luciferases / Q000378:metabolism; D011401:Promoter Regions, Genetic; D051381:Rats; D021541:Sea Bream / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection / Q000662:veterinary
chemical_list: D016207:Cytokines; D029941:Fish Proteins; D005938:Glucocorticoids; D015850:Interleukin-6; D008070:Lipopolysaccharides; D008156:Luciferases