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Identification of a novel tumor necrosis factor alpha-responsive region in the NCF2 promoter.
J Leukoc Biol. 2005 Feb; 77(2):267-78.JL

Abstract

The phagocyte reduced nicotinamide adenine dinucleotide phosphate oxidase is a multiprotein enzyme that catalyzes the production of microbicidal oxidants. Although oxidase assembly involves association of several membrane and cytosolic oxidase proteins, one of the cytosolic cofactors, p67phox, appears to play a more prominent role in final activation of the enzyme complex. Based on the importance of p67phox, we investigated transcriptional regulation of the p67phox gene [neutrophil cytosolic factor 2 (NCF2)] and demonstrated previously that activator protein-1 (AP-1) was essential for basal transcriptional activity. As p67phox can be up-regulated by tumor necrosis factor alpha (TNF-alpha), which activates AP-1, we hypothesized that TNF-alpha might regulate NCF2transcription via AP-1. In support of this hypothesis, we show here that NCF2 promoter-reporter constructs are up-regulated by TNF-alpha but only when AP-1 factors were coexpressed. Consistent with this observation, we also demonstrate that NCF2 mRNA and p67phox protein are up-regulated by TNF-alpha in various myeloid cell lines as well as in human monocytes. It was surprising that mutagenesis of the AP-1 site in NCF2 promoter constructs did not eliminate TNF-alpha induction, suggesting additional elements were involved in this response and that AP-1 might play a more indirect role. Indeed, we used NCF2 promoter-deletion constructs to map a novel TNF-alpha-responsive region (TRR) located between -56 and -16 bp upstream of the translational start site and demonstrated its importance in vivo using transcription factor decoy analysis. Furthermore, DNase footprinting verified specific binding of factor(s) to the TRR with AP-1 binding indirectly to this region. Thus, we have identified a novel NCF2 promoter/enhancer domain, which is essential for TNF-alpha-induced up-regulation of p67phox.

Authors+Show Affiliations

Department of Veterinary Molecular Biology, Montana State University, Bozeman, MT 59717, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

15513967

Citation

Gauss, Katherine A., et al. "Identification of a Novel Tumor Necrosis Factor Alpha-responsive Region in the NCF2 Promoter." Journal of Leukocyte Biology, vol. 77, no. 2, 2005, pp. 267-78.
Gauss KA, Bunger PL, Larson TC, et al. Identification of a novel tumor necrosis factor alpha-responsive region in the NCF2 promoter. J Leukoc Biol. 2005;77(2):267-78.
Gauss, K. A., Bunger, P. L., Larson, T. C., Young, C. J., Nelson-Overton, L. K., Siemsen, D. W., & Quinn, M. T. (2005). Identification of a novel tumor necrosis factor alpha-responsive region in the NCF2 promoter. Journal of Leukocyte Biology, 77(2), 267-78.
Gauss KA, et al. Identification of a Novel Tumor Necrosis Factor Alpha-responsive Region in the NCF2 Promoter. J Leukoc Biol. 2005;77(2):267-78. PubMed PMID: 15513967.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Identification of a novel tumor necrosis factor alpha-responsive region in the NCF2 promoter. AU - Gauss,Katherine A, AU - Bunger,Peggy L, AU - Larson,Trina C, AU - Young,Catherine J, AU - Nelson-Overton,Laura K, AU - Siemsen,Daniel W, AU - Quinn,Mark T, Y1 - 2004/10/28/ PY - 2004/10/30/pubmed PY - 2005/3/16/medline PY - 2004/10/30/entrez SP - 267 EP - 78 JF - Journal of leukocyte biology JO - J Leukoc Biol VL - 77 IS - 2 N2 - The phagocyte reduced nicotinamide adenine dinucleotide phosphate oxidase is a multiprotein enzyme that catalyzes the production of microbicidal oxidants. Although oxidase assembly involves association of several membrane and cytosolic oxidase proteins, one of the cytosolic cofactors, p67phox, appears to play a more prominent role in final activation of the enzyme complex. Based on the importance of p67phox, we investigated transcriptional regulation of the p67phox gene [neutrophil cytosolic factor 2 (NCF2)] and demonstrated previously that activator protein-1 (AP-1) was essential for basal transcriptional activity. As p67phox can be up-regulated by tumor necrosis factor alpha (TNF-alpha), which activates AP-1, we hypothesized that TNF-alpha might regulate NCF2transcription via AP-1. In support of this hypothesis, we show here that NCF2 promoter-reporter constructs are up-regulated by TNF-alpha but only when AP-1 factors were coexpressed. Consistent with this observation, we also demonstrate that NCF2 mRNA and p67phox protein are up-regulated by TNF-alpha in various myeloid cell lines as well as in human monocytes. It was surprising that mutagenesis of the AP-1 site in NCF2 promoter constructs did not eliminate TNF-alpha induction, suggesting additional elements were involved in this response and that AP-1 might play a more indirect role. Indeed, we used NCF2 promoter-deletion constructs to map a novel TNF-alpha-responsive region (TRR) located between -56 and -16 bp upstream of the translational start site and demonstrated its importance in vivo using transcription factor decoy analysis. Furthermore, DNase footprinting verified specific binding of factor(s) to the TRR with AP-1 binding indirectly to this region. Thus, we have identified a novel NCF2 promoter/enhancer domain, which is essential for TNF-alpha-induced up-regulation of p67phox. SN - 0741-5400 UR - https://www.unboundmedicine.com/medline/citation/15513967/Identification_of_a_novel_tumor_necrosis_factor_alpha_responsive_region_in_the_NCF2_promoter_ L2 - https://doi.org/10.1189/jlb.0604329 DB - PRIME DP - Unbound Medicine ER -