The proximal promoter region is essential for lipopolysaccharide induction and cyclic AMP inhibition of mouse tumor necrosis factor-alpha.
J Interferon Cytokine Res
; 22(5): 539-48, 2002 May.
Article
en En
| MEDLINE
| ID: mdl-12060492
Bacterial lipopolysaccharide (LPS) induces a rapid and transient increase in transcription of the tumor necrosis factor-alpha (TNF-alpha) gene in cells of monocyte/macrophage lineage. This study examines the role of potential regulatory elements within the proximal promoter region of the mouse TNF-alpha gene in LPS induction and cyclic AMP (cAMP)-mediated inhibition of TNF-alpha in RAW 264.7 murine macrophage-like cells. Transfection of proximal promoter chloramphenicol acetyltransferase (CAT) reporter constructs demonstrated that this region is LPS inducible in murine RAW 264.7 cells, with a 5.9-fold increase over nonstimulated transfectants. Site-specific mutations of the ETS, activated protein-1 (AP-1)/cAMP-responsive element (CRE)-like, or NF-kappaB-like motifs within this region caused a reduction in the LPS response by 52%, 46%, and 51%, respectively. LPS induction of the proximal promoter-CAT reporter construct was reduced by >40% by the addition of 8-bromo-cAMP (8-Br-cAMP). To determine the role of the proximal promoter region in the context of the entire TNF-alpha gene, we produced a hemagglutinin (HA)-tagged genomic TNF-alpha construct that contains a deletion of the proximal promoter region. Transfection of this construct into RAW 264.7 cells demonstrated a decrease in LPS-induced transcripts as well as a lack of response to cAMP. This suggested an essential role for this regulatory region in LPS-induced activation and cAMP inhibition of mouse TNF-alpha gene transcription in murine macrophages.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Lipopolisacáridos
/
Regiones Promotoras Genéticas
/
Factor de Necrosis Tumoral alfa
/
AMP Cíclico
Límite:
Animals
Idioma:
En
Revista:
J Interferon Cytokine Res
Asunto de la revista:
ALERGIA E IMUNOLOGIA
Año:
2002
Tipo del documento:
Article
País de afiliación:
Estados Unidos
Pais de publicación:
Estados Unidos