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The Journal of Immunology, 2003, 171: 1750-1757.
Copyright © 2003 by The American Association of Immunologists

RNA Interference Shows Critical Requirement for NF-{kappa}B p50 in the Production of IL-12 by Human Dendritic Cells1

Diego Laderach*, Daniel Compagno{dagger}, Olivier Danos{dagger}, William Vainchenker* and Anne Galy2,*,{dagger}

* Institut National de la Santé et de la Recherche Médicale Unité 362, Institut Gustave Roussy, Villejuif, France; and {dagger} Généthon-Centre National de la Recherche Scientifique Unité Mixte de Recherche 8115, Evry, France

Specific NF-{kappa}B/Rel proteins regulate murine dendritic cell (DC) survival, differentiation, and activation, but little is known of their role in human cells because of limited loss-of-function analyses. RNA interference (RNAi) is a mechanism to effectively silence gene expression via sequence-specific double-stranded small interfering RNAs (siRNAs). RNAi was used to assess the role of the p50 (NF-{kappa}B1) protein in the maturation and activation of cultured human monocyte-derived DC (MoDC). Transfection of cultured MoDC with siRNAs reduced p50 mRNA and protein levels in a specific, dose-dependent, and time-dependent manner. Basal or maturation-induced expression of HLA-DR and costimulatory molecules were not affected, whereas transcription of the IL-12 p40 gene and the secretion of IL-12{alpha}{beta} were reduced. Such MoDC induced less IFN-{gamma} production by T cells in MLR. This is the first report of RNAi-induced phenotype in human primary DC with a method that caused no measurable toxicity or type-I IFN response. siRNAs appear useful for the study of signaling pathways in immune cells, revealing a pivotal requirement for p50 in MoDC for IL-12 production and induction of optimal type-1 immune responses.




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