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Cellular events mediated by lipopolysaccharide-stimulated toll-like receptor 4. MD-2 is required for activation of mitogen-activated protein kinases and Elk-1.
Journal of Biological Chemistry 2000 July 8
Lipopolysaccharide (LPS) stimulates multiple signaling events, including nuclear factor-kappaB (NF-kappaB) activity and the mitogen-activated protein (MAP) kinases, ERK, JNK, and p38 in LPS-responsive cells, resulting in transcriptional activation and cytokine generation. LPS-induced signaling via toll-like receptor 4 (TLR4) results in the activation of the transcription factor NF-kappaB. Since LPS activates other signaling cascades in responsive cells, the objective of this study was to determine whether such events are mediated by TLR4 in response to LPS. We generated human embryonic kidney cells (HEK293) that stably express TLR4 (HEK-TLR4) and examined their responsiveness to LPS by measuring NF-kappaB activity and production of interleukin-8 (IL-8). A trans-reporting system was used to measure the activity of Elk-1, an ETS-domain transcription factor targeted by MAP kinase pathways. LPS stimulated NF-kappaB reporter activity and IL-8 production but not Elk-1 activity in HEK-TLR4 cells. When MD-2, a protein associated with the extracellular domain of TLR4, was expressed in these cells, there was a marked increase in Elk-1 activity as well as ERK, JNK, and p38 MAP kinase phosphorylation in response to LPS. TLR4-mediated NF-kappaB reporter activity and IL-8 production was enhanced by the expression of MD-2. This study demonstrates that expression of both TLR4 and MD-2 is required for LPS to activate or augment the MAP kinase pathways, Elk-1 stimulation, and IL-8 generation.
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