title: Cloning and characterization of the porcine IL-10 promoter.
abstract: Interleukin 10 (IL-10) is an anti-inflammatory and immunosuppressive cytokine that plays an important role in regulating the immune response. Therefore, understanding how IL-10 is regulated is important. The regulatory elements have been well studied in human and mouse promoters and several transcription factors have been showed to be involved in IL-10 transcription. In our study, a 1.5 kb fragment of the 5' flanking region of IL-10 gene was cloned and functionally characterized. Several putative regulatory elements including IRF, AP-1, Sp1, C/EBP, and STAT binding sites were found in the porcine IL-10 (pIL-10) promoter. The pIL-10 promoter deletion mutants were analyzed for their ability to direct luciferase expression in a porcine macrophage cell line (CRL 2843), human gastric carcinoma cell lines with or without Epstein-Barr virus (EBV), AGS-EBV and AGS cell lines. Our data showed that the minimal active pIL-10 promoter region was from -605 to +19, with the inducible activity requiring only one key DNA element, the Sp1 binding site (-398 to -393) upstream of the IL-10 gene starting point in both LPS-stimulated CRL 2843 and AGS-EBV cells. Moreover, our results suggested that the two IRF binding sites (-950 to -942 and -662 to -640) may have a positive role in the activation of the pIL-10 promoter in AGS-EBV cells, but not in LPS-stimulated CRL 2843 cells. These data implicate that the cloned porcine IL-10 promoter could be used to explore the molecular mechanisms underlying the regulation of IL10 production in pigs.
authors.affiliation: State key laboratory of Agrobiotechnology, Department of Microbiology and Immunology, College of Biological Science, China Agricultural University, Beijing 100193, China.
authors.forename: Rong
authors.initials: R
authors.lastname: Quan
authors.forename: Yi
authors.initials: Y
authors.lastname: Fu
authors.forename: Weiyong
authors.initials: W
authors.lastname: He
authors.forename: Wen-Hai
authors.initials: WH
authors.lastname: Feng
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D045744:Cell Line, Tumor; D003001:Cloning, Molecular / Q000379:methods; D007136:Immunoglobulins / Q000235:genetics / Q000276:immunology; D050835:Interferon Regulatory Factors / Q000235:genetics / Q000276:immunology; D016753:Interleukin-10 / Q000235:genetics* / Q000276:immunology*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016133:Polymerase Chain Reaction / Q000662:veterinary; D011401:Promoter Regions, Genetic; D016415:Sequence Alignment; D013552:Swine / Q000235:genetics* / Q000276:immunology
chemical_list: D007136:Immunoglobulins; D050835:Interferon Regulatory Factors; C090698:SP1 antigen; D016753:Interleukin-10