A distal regulatory region is required for constitutive and IFN-beta-induced expression of murine TLR9 gene

TLR9 is critical for the recognition of unmethylated CpG DNA in innate immunity. Accumulating evidence suggests distinct patterns of TLR9 expression in various types of cells. However, the molecular mechanism of TLR9 expression has received little attention. In the present study, we demonstrate that...

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Bibliographic Details
Published in:Journal of Immunology 2005-12, Vol.175 (11), p.7407-7418
Main Authors: Guo, Zhu, Garg, Sanjay, Hill, Karen M, Jayashankar, Lakshmi, Mooney, Myesha R, Hoelscher, Mary, Katz, Jacqueline M, Boss, Jeremy M, Sambhara, Suryaprakash
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Base Sequence
Cell Line
DNA Footprinting
DNA Primers
Electrophoretic Mobility Shift Assay
Gene Expression
Gene Expression Regulation - immunology
Interferon Regulatory Factor-1 - genetics
Interferon Regulatory Factor-1 - immunology
Interferon Regulatory Factor-2 - genetics
Interferon Regulatory Factor-2 - immunology
Interferon-beta - immunology
Interferon-beta - metabolism
Macrophages - immunology
Mice
Molecular Sequence Data
Promoter Regions, Genetic
Proto-Oncogene Proteins c-fos - genetics
Proto-Oncogene Proteins c-fos - immunology
Proto-Oncogene Proteins c-jun - genetics
Proto-Oncogene Proteins c-jun - immunology
Regulatory Sequences, Nucleic Acid - immunology
Reverse Transcriptase Polymerase Chain Reaction
RNA Interference
STAT1 Transcription Factor - genetics
STAT1 Transcription Factor - immunology
Toll-Like Receptor 2 - genetics
Toll-Like Receptor 2 - immunology
Toll-Like Receptor 9 - genetics
Transcription Factor AP-1 - genetics
Transcription Factor AP-1 - immunology
Transcription, Genetic
Transfection
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Summary:TLR9 is critical for the recognition of unmethylated CpG DNA in innate immunity. Accumulating evidence suggests distinct patterns of TLR9 expression in various types of cells. However, the molecular mechanism of TLR9 expression has received little attention. In the present study, we demonstrate that transcription of murine TLR9 is induced by IFN-beta in peritoneal macrophages and a murine macrophage cell line RAW264.7. TLR9 is regulated through two cis-acting regions, a distal regulatory region (DRR) and a proximal promoter region (PPR), which are separated by approximately 2.3 kbp of DNA. Two IFN-stimulated response element/IFN regulatory factor-element (ISRE/IRF-E) sites, ISRE/IRF-E1 and ISRE/IRF-E2, at the DRR and one AP-1 site at the PPR are required for constitutive expression of TLR9, while only the ISRE/IRF-E1 motif is essential for IFN-beta induction. In vivo genomic footprint assays revealed constitutive factor occupancy at the DRR and the PPR and an IFN-beta-induced occupancy only at the DRR. IRF-2 constitutively binds to the two ISRE/IRF-E sites at the DRR, while IRF-1 and STAT1 are induced to bind to the two ISRE/IRF-E sites and the ISRE/IRF-E1, respectively, only after IFN-beta treatment. AP-1 subunits, c-Jun and c-Fos, were responsible for the constitutive occupancy at the proximal region. Induction of TLR9 by IFN-beta was absent in STAT1-/- macrophages, while the level of TLR9 induction was decreased in IRF-1-/- cells. This study illustrates the crucial roles for AP-1, IRF-1, IRF-2, and STAT1 in the regulation of murine TLR9 expression.
ISSN:0022-1767
1550-6606
1365-2567
DOI:10.4049/jimmunol.175.11.7407