title: Fish IRF3 up-regulates the transcriptional level of IRF1, IRF2, IRF3 and IRF7 in CIK cells.
abstract: Interferon Regulatory Factors (IRFs) belong to a family of transcription factor involved in transcriptional regulation of type I IFN and IFN-stimulated genes (ISG) in cells. In the present study, an IRF3 full-length cDNA (termed CiIRF3, JX999055) and its promoter sequence were cloned by homology cloning strategy and genome walking from grass carp (Ctenopharyngodon idella). The full-length cDNA sequence of CiIRF3 is comprised of a 5'UTR (195 bp), a 3'UTR (269 bp) and a largest open reading frame (ORF) of 1377 bp encoding a polypeptide of 458 amino acids. CiIRF3 has a conservative DNA-binding domain (DBD) at N-terminal and a relatively conserved interferon regulatory factors association domain (IAD). Phylogenetic tree analysis indicated that CiIRF3 gathers together with other IRF-3 from higher vertebrates in the same branch. The promoter sequence of CiIRF3 (596 bp) consists of three IRF-E, a C/EBP beta, a NF-kappa B and a TATA-BOX. CiIRF3 was constitutively expressed at low level in different grass carp tissues but was rapidly up-regulated with Poly I:C stimulation. To study the molecular mechanism of CiIRF3 regulating the transcription of IRFs, CiIRF3 was expressed in Escherichia coli BL21 and purified by affinity chromatography with the Ni-NTA His-Bind Resin. Gel mobility shift assays revealed the affinity of CiIRF3 protein with promoters of CiIRF1, CiIRF2, CiIRF3 and CiIRF7 respectively. Then, CIK cells were co-transfected with pcDNA3.1-CiIRF3, pGL3-promotor (pGL3-CiIRF1, pGL3-CiIRF2, pGL3-CiIRF3, pGL3-CiIRF7) and luciferase reporter vector respectively. The cotransfection experiment showed that CiIRF3 increased the promoter activity of CiIRF1, CiIRF2, CiIRF3 and CiIRF7. Furthermore, overexpression of CiIRF3 in CIK cells also up-regulated the expressions of CiIRF1, CiIRF2, CiIRF3 and CiIRF7. So, CiIRF3 can improve the transcriptional level of CiIRF1, CiIRF2, CiIRF3 and CiIRF7.
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China.
authors.forename: Xiaowen
authors.initials: X
authors.lastname: Xu
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China; Infectious Diseases Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, China.
authors.forename: Qinan
authors.initials: Q
authors.lastname: Lai
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China.
authors.forename: Meihui
authors.initials: M
authors.lastname: Gu
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China.
authors.forename: Dan
authors.initials: D
authors.lastname: Liu
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China.
authors.forename: Qunhao
authors.initials: Q
authors.lastname: Hou
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China.
authors.forename: Xiancheng
authors.initials: X
authors.lastname: Liu
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China.
authors.forename: Yichuan
authors.initials: Y
authors.lastname: Mi
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China.
authors.forename: Zhicheng
authors.initials: Z
authors.lastname: Sun
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China.
authors.forename: Haizhou
authors.initials: H
authors.lastname: Wang
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China.
authors.forename: Gang
authors.initials: G
authors.lastname: Lin
authors.affiliation: Department of Bioscience, College of Life Science, Nanchang University, Nanchang 330031, China. Electronic address: hucy2008@163.com.
authors.forename: Chengyu
authors.initials: C
authors.lastname: Hu
mesh_terms: D000818:Animals; D001483:Base Sequence; D002347:Carps / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D002460:Cell Line; D004926:Escherichia coli / Q000235:genetics; D029941:Fish Proteins / Q000235:genetics* / Q000378:metabolism; D050835:Interferon Regulatory Factors / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D011070:Poly I-C / Q000494:pharmacology; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015533:Transcriptional Activation
chemical_list: D029941:Fish Proteins; D050835:Interferon Regulatory Factors; D011994:Recombinant Proteins; D011070:Poly I-C
keywords: Fish; IRF3; ISG; Promoter; Transcriptional control