Functional Interaction of HIF-1 and NF-kappaB Increasing the Transcriptional Activation of TNF-alpha Gene in Monocytes.
- Author:
Min Jeong PARK
1
;
Sun Min LEE
;
Soon Jung OK
;
Hye Rim KIM
;
Hyung Hoi KIM
;
Jae Hun CHEONG
Author Information
1. Department of Molecular Biology, College of Natural Sciences, Pusan National University, Busan, Korea. molecule85@pusan.ac.kr
- Publication Type:Original Article
- Keywords:
HIF-1;
NF-kappaB;
Hypoxia;
TNF-alpha;
Transactivation
- MeSH:
Anoxia;
Binding Sites;
Chromatin;
Chromatin Immunoprecipitation;
Electroporation;
Enhancer Elements, Genetic;
Gene Expression;
Humans;
Immunoprecipitation;
Inflammation;
Leukemia;
Macrophages;
Monocytes;
NF-kappa B;
Proteins;
Transcription Factors;
Transcriptional Activation;
Tumor Necrosis Factor-alpha;
Two-Hybrid System Techniques;
U937 Cells
- From:Korean Journal of Blood Transfusion
2013;24(1):21-32
- CountryRepublic of Korea
- Language:English
-
Abstract:
BACKGROUND: Tumor necrosis factor alpha (TNF-alpha) is a pleiotropic cytokine fulfilling a broad variety of immunoregulatory functions. Monocytes and macrophages play a pivotal role in inflammation and immune regulation. NF-kappaB and HIF-1 are known to increase expression of the TNF-alpha gene in a separate way. METHODS: Human monocytic leukemia, U937 cells, were transfected using the standard electroporation method for intracellular expression of NF-kappaB and HIF-1. We performed analysis using the mammalian two-hybrid assay and co-immunoprecipitation assay for detection of protein interaction of both proteins. In addition, chromatin immunoprecipitation analysis was performed for examination of NF-kappaB and HIF-1 binding on the TNF-alpha gene promoter. RESULTS: Here we show that NF-kappaB and HIF-1 cooperatively induced an increase in expression of the TNF-alpha gene dependent on promoter activity by the direct protein interaction of these two transcription factors. Hypoxia signaling induced marked enhancement of the transactivation of TNF-alpha promoter by HIF-1 and NF-kappaB. A tandem NF-kappaB/HIF-1 binding site was identified within the TNF-alpha promoter, which acted as a strong enhancer element. Physical association of the Rel domain of NF-kappaB and the N-TD domain of HIF-1 was required. Hypoxia treatment also resulted in a significant increase in the protein interaction of NF-kappaB and HIF-1 in vivo. Both transcription factors were recruited on the chromatin TNF-alpha promoter dependent on hypoxia stimuli. CONCLUSION: The results of this study indicate that a variety of extracellular signals for activation of TNF-alpha gene expression might converge on the transcriptional regulation through the NF-kappaB/HIF-1 signaling pathway.
