IFI-16 GENE ENCODES A NUCLEAR-PROTEIN WHOSE EXPRESSION IS INDUCED BY INTERFERONS IN HUMAN MYELOID-LEUKEMIA CELL-LINES

We have characterized the induction of mRNA and protein products of th e human IFI 16 gene in response to IFN-gamma, IFN-alpha, and IFN-beta( 2) (IL-6). We demonstrate that the IFI 16 gene product is a novel nucl eoprotein expressed in association with the differentiation of myeloid precursor cell lines. In Northern blots, IFI 16 mRNA was increased si milar to 25-fold above barely detectable levels in unstimulated promye locytic HL-60 cells, in response to IFN-gamma. Other myeloid cell line s, U937 and K562, also demonstrated a marked IFN-gamma-inducibility of IFI 16 mRNA. However, all three cell lines were far less responsive t o IFN-alpha, and there was no response to IL-6. By comparison, a panel of T and B cell lines demonstrated high constitutive expression of IF I 16 mRNA that was not regulated by these cytokines. Culture of HL-60 cells in medium containing dimethylsulfoxide, retinoic acid, and 1,25 dihydroxyvitamin D-3, agents that stimulate the differentiation of HL- 60 along myeloid pathways, also caused the induction of IFI 16 mRNA. T o characterize the protein product of IN 16, a monoclonal antibody was raised against a recombinant bacterial protein comprising the amino t erminal 159 amino acids of IFI 16 fused to glutathione S-transferase. The antibody, designated 1G7, was used in Western blotting to demonstr ate the strong induction of a cluster of proteins of 85-95 kDa in the nuclear extracts of IFN-gamma-treated HL-60. The nuclear localization of IFI 16 antigen was confirmed by immunohistochemical staining of HL- 60 cells treated with IFN-gamma, dimethylsulfoxide, and retinoic acid. IFI 16 was also detected in the nuclei of monocytes, neutrophils, and lymphocytes in normal peripheral blood. Database comparisons of the I FI 16 amino acid sequence revealed 51% identity with the recently clon ed myeloid cell nuclear differentiation antigen (MNDA), and extensive similarity to protein products of the Gene 200 cluster of IFN-inducibl e genes, Ifi 202 and Ifi 204. The amino terminal domain of IFI 16 enco des a putative nuclear localization signal, (124)PGAQKRKK, which is st rongly conserved in MNDA and 204. Nuclear IFI 16 was able to bind doub le-stranded DNA in vitro and exhibited a similar elution profile from DNA-cellulose as previously observed for MNDA and 204. Therefore, IFI 16 and MNDA are members of a novel family of human DNA-binding protein s whose expression is associated with myeloid cell differentiation ind uced by cytokines and chemical agents. (C) 1995 Wiley-Liss, Inc.