Mechanisms regulating expression of the tumor necrosis factor-related light gene. Role of calcium-signaling pathway in the transcriptional control.
LIGHT (TNFSF14) is a newly identified tumor necrosis factor superfamily member involved in the regulation of immune responses by control of activation, maturation, and survival of immune effector cells. Despite the immunological relevance of the LIGHT protein, little knowledge is available as to how light gene expression is regulated. In T-lymphocytes, most LIGHT surface expression and transcript accumulation occurs after T cell activation. In this study, we have shown that these events are blocked at the transcriptional level by cyclosporin A, an immuno-suppressive drug. Besides, we identified a role for Ca2+ -signaling pathways and NFAT transcription factors in T cell activation-induced LIGHT expression. To further investigate this process, we have identified, cloned, and characterized a 2.1-kilobase 5'-flanking DNA genomic fragment from the human light gene. We have shown the transcriptional activity of the herein-identified minimal 5' regulatory region of human light gene parallels the endogenous expression of light in T cells. Moreover, we demonstrated that induced LIGHT promoter activity can be equally blocked by cyclosporin A treatment or dominant negative NFAT overexpression and further identified by site-directed mutagenesis and electrophoretic mobility supershift analysis of a NFAT transcription factor binding site within the human light minimal promoter. Finally, Sp1 and Ets1 binding sites were identified and shown to regulate light basal promoter activity. Thus, the present study establishes a molecular basis to further understand the mechanisms governing human light gene expression and, consequently, could potentially lead to novel therapeutic manipulations that control the signaling cascade, resulting in LIGHT production in conditions characterized by immunopathologic activation of T cells.
Castellano
Remy
R
Institut de Cancérologie et d'Immunologie de Marseille, Université de la Méditerranée, INSERM, Unite 119, 13009 Marseille, France.
Van Lint
Carine
C
Peri
Valentine
V
Veithen
Emmanuelle
E
Morel
Yannis
Y
Costello
Regis
R
Olive
Daniel
D
Collette
Yves
Y
D001483:Base Sequence; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D016572:Cyclosporine / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects / Q000276:immunology; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology; D019169:Jurkat Cells; D008213:Lymphocyte Activation; D008565:Membrane Proteins / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D013601:T-Lymphocytes / Q000276:immunology*; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects / Q000276:immunology; D053326:Tumor Necrosis Factor Ligand Superfamily Member 14; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics* / Q000276:immunology
D004268:DNA-Binding Proteins; D007166:Immunosuppressive Agents; D008565:Membrane Proteins; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D011994:Recombinant Proteins; C505914:TNFSF14 protein, human; D014157:Transcription Factors; D053326:Tumor Necrosis Factor Ligand Superfamily Member 14; D014409:Tumor Necrosis Factor-alpha; D016572:Cyclosporine