Abstract
Interferon (IFN)-γ-induced expression of indoleamine 2,3-dioxygenase (IDO) gene is implicated in the antimicrobial and antiproliferative effects of IFN-γ in cell cultures. Earlier studies identified a 96 base pair (bp) regulatory region upstream of the IDO gene that conferred IFN-γ response to the chloroamphenicol acetyltransferase (CAT) gene linked to herpesvirus thymidine kinase promoter. The IFN-γ-responsive region was further narrowed to a 67 bp fragment by 3′ deletion. This 67 bp fragment contains several sequence elements of potential interest, including a 14 bp sequence homologous to the ISRE sequence found in IFN-α-inducible genes and two palindromic sequences (PE I and PE II) homologous to the GAS sequence identified in IFN-γ-mducible genes. Site-directed mutagenesis studies showed that IFN-γ-induced expression of IDO-CAT constructs involved cooperation between two elements: the ISRE homolog and the PE II (but not PE I). Either element alone with its flanking sequence was inadequate in conferring an IFN-γ response to CAT reporter gene. Two IFN-γ-regulated protein factors interacting with these two elements were identified. The factor binding to the ISRE region was induced with a slower kinetics, required new protein synthesis, and reacted with antibodies to IRF-1. The factor interacting with the PE II region appeared rapidly after treatment with IFN-γ independently of new protein synthesis, and its binding to DNA probe was blocked by antibodies to p91 factor, reported to bind to GAS element. The results indicate that the IFN-γ-inducible expression of the human IDO gene differs from that of other IFN-γ-inducible genes with respect to a complete dependence on two control elements, an ISRE and a GAS element (PE II), which interact with IFN-γ-regulated factors IRF-1 and p91. The involvement of IRF-1 explains the dependence on new protein synthesis for IFN-γ-mediated induction of the IDO gene.
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