CD200 plays an important role in prevention of graft rejection, autoimmune diseases and spontaneous abortion by delivering an immunoregulatory signal through interaction with its receptor. It also plays a role in regulating tumor immunity. We previously documented evidence for C/EBPβ as being important in the regulation of constitutive expression of CD200. However, the molecular mechanism(s) controlling inducible expression of CD200 are yet to be explored. Here we address the regulated expression of human CD200 by T cells in response to Con A, IFN-γ or/and TNF-α. A prominent DNase I hypersensitivity site (DHS) was localized ∼5.4 kb upstream of the major transcriptional start site. Four cis-elements were identified within this genomic region: one nuclear factor-κB (NF-κB) site, one IFN-γ activation site (GAS) element and two IFN-stimulated response element (ISRE) for binding of interferon-regulatory factors (IRFs), respectively. Mutation of the NF-κB site, GAS or one but not the other of ISREs dramatically reduced the luciferase activity. These findings were further confirmed by chromatin immunoprecipitation (ChIP) assays using antibodies against NF-κB p65, STAT1α, and IRF-1. All the above findings suggest that IFN-γ and TNF-α induce CD200 expression through a 5′ upstream enhancer and that NF-κB, STAT1 and IRF-1 play pivotal roles in this process.