Characterization of an activation protein-1-binding site in the murine interleukin-12 p40 promoter. Demonstration of novel functional elements by a reductionist approach.
Interleukin (IL)-12 is a heterodimeric cytokine produced by macrophages in response to intracellular pathogens and provides an obligatory signal for the differentiation of T-helper-1 cells. We previously reported an analysis of the IL-12 p40 promoter in RAW264.7 macrophages. Multiple control elements were involved in activation of transcription by bacterial products. A critical control element, located between -96 and -88, interacts with C/EBP family members. In this study, using a strategy to demonstrate functional activity in a minimal promoter context, three novel cis-acting elements are found to have an important role in IL-12 p40 promoter activation by lipopolysaccharide. One of these elements is characterized in detail. Mutations from -79 to -74 in the murine IL-12 p40 promoter significantly reduce lipopolysaccharide-induced promoter activity. Electrophoretic mobility shift assays demonstrate binding of AP-1 family members to this region. Spacing between the C/EBP and AP-1 site is important for promoter activation, suggesting cooperativity between these elements. c-Jun and a mutant c-Jun molecule activate the IL-12 p40 promoter and synergistically activate the promoter when co-expressed with C/EBPbeta. Finally, this region of the promoter is demonstrated to be a target for mitogen-activated protein kinase and toll-like receptor signaling pathways.
Zhu
C
C
Immunobiology Center, The Mount Sinai School of Medicine, New York, New York 10029-6574, USA.
Gagnidze
K
K
Gemberling
J H
JH
Plevy
S E
SE
D000818:Animals; D018664:Interleukin-12 / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D011401:Promoter Regions, Genetic; D015398:Signal Transduction; D018808:Transcription Factor AP-1 / Q000235:genetics* / Q000378:metabolism
D018808:Transcription Factor AP-1; D018664:Interleukin-12