Cytokine-responsive induction of SAF-1 activity is mediated by a mitogen-activated protein kinase signaling pathway.
SAF-1, a zinc finger transcription factor, is activated by a number of inflammatory agents, including interleukin-1 (IL-1) and IL-6. It is involved in the cytokine-mediated transcriptional induction of serum amyloid A, an acute-phase plasma protein that is associated with the pathogenesis of reactive amyloidosis, rheumatoid arthritis, and atherosclerosis. Here, we show that the mitogen-activated protein (MAP) kinase signaling pathway regulates cytokine-mediated induction of the DNA-binding activity and transactivation potential of SAF-1. Phosphorylation of endogenous SAF-1 in response to IL-1 and IL-6 was markedly inhibited by the addition of MAP kinase inhibitors. Consistent with this finding, we show that a consensus MAP kinase phosphorylation site, PPTP, within SAF-1 could be phosphorylated by MAP kinase in vitro. To analyze the contribution of MAP kinase in the activation of SAF-1, we prepared two independent mutant proteins in which the threonine residue of the PPTP motif was altered to either valine or alanine. These mutant proteins lost the ability to be phosphorylated by MAP kinase both in vivo and in vitro and exhibited a significantly reduced ability to promote expression of the SAF-1-regulated promoter. While the DNA-binding activity of wild-type SAF-1 protein was markedly increased upon phosphorylation with MAP kinase, no such increase could be detected with the mutant SAF-1 proteins. Further analysis with the GAL-4 reporter system showed that mutation of the MAP kinase phosphorylation site considerably lowers the transactivation potential of SAF-1. Together, these results show that activation of SAF-1 in response to IL-1 and -6 is mediated via MAP kinase-regulated phosphorylation.
Ray
Alpana
A
Department of Veterinary Pathobiology, University of Missouri, Columbia, Missouri 65211, USA. Rayal@missouri.edu
Yu
Guang-Yao
GY
Ray
Bimal K
BK
D020816:Amino Acid Motifs; D000818:Animals; D002460:Cell Line; D016207:Cytokines / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000502:physiology*; D017930:Genes, Reporter; D006801:Humans; D007375:Interleukin-1 / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation / Q000502:physiology; D016335:Zinc Fingers
D016207:Cytokines; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D007375:Interleukin-1; D015850:Interleukin-6; D009687:Nuclear Proteins; D014157:Transcription Factors; C076035:c-MYC-associated zinc finger protein; D020928:Mitogen-Activated Protein Kinases