Identification of target genes and a unique cis element regulated by IRF-8 in developing macrophages

• Published in: Blood Vol. 106; no. 6; pp. 1938 - 1947
• Main Authors: Tamura, Tomohiko, Thotakura, Pratima, Tanaka, Tetsuya S., Ko, Minoru S.H., Ozato, Keiko
• Format: Journal Article
• Language: English
• Published: Washington, DC Elsevier Inc 15-09-2005; The Americain Society of Hematology; 2005 by The American Society of Hematology
• Subjects: Animals; Base Sequence; Binding Sites; Biological and medical sciences; Cathepsin C - genetics; Cell Differentiation - genetics; Cell differentiation, maturation, development, hematopoiesis; Cell Line; Cell physiology; Chromatin Immunoprecipitation; Consensus Sequence; Cystatin C; Cystatins - genetics; Enhancer Elements, Genetic - genetics; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation; Genes, Reporter; Hematopoiesis; Interferon Regulatory Factors; Macrophages - cytology; Macrophages - metabolism; Mice; Molecular and cellular biology; Oligonucleotide Array Sequence Analysis; Promoter Regions, Genetic; Repressor Proteins - physiology; Transcription, Genetic; Enzyme; DNA chip; Interferon consensus sequence binding protein; Gene expression; Peptidases; Signal transduction; Cathepsin; Transcription factor PU.1; Dipeptidyl-peptidase I; Gene; Hematopoiesis; Cystatin C; Hydrolases; Genetics; Myelopoiesis; Transcription factor; Macrophage
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood-2005-01-0080

Interferon regulatory factor-8 (IRF-8)/interferon consensus sequence–binding protein (ICSBP) is a transcription factor that controls myeloid-cell development. Microarray gene expression analysis of Irf-8-/- myeloid progenitor cells expressing an IRF-8/estrogen receptor chimera (which differentiate into macrophages after addition of estradiol) was used to identify 69 genes altered by IRF-8 during early differentiation (62 up-regulated and 7 down-regulated). Among them, 4 lysosomal/endosomal enzyme-related genes (cystatin C, cathepsin C, lysozyme, and prosaposin) did not require de novo protein synthesis for induction, suggesting that they were direct targets of IRF-8. We developed a reporter assay system employing a self-inactivating retrovirus and analyzed the cystatin C and cathepsin C promoters. We found that a unique cis element mediates IRF-8–induced activation of both promoters. Similar elements were also found in other IRF-8 target genes with a consensus sequence (GAAANN[N]GGAA) comprising a core IRF-binding motif and an Ets-binding motif; this sequence is similar but distinct from the previously reported Ets/IRF composite element. Chromatin immunoprecipitation assays demonstrated that IRF-8 and the PU.1 Ets transcription factor bind to this element in vivo. Collectively, these data indicate that IRF-8 stimulates transcription of target genes through a novel cis element to specify macrophage differentiation.