Abstract
We have previously demonstrated that the antiapoptotic gene mcl-1 is activated by interleukin-3 (IL-3) in Ba/F3 pro-B cells through two promoter elements designated the CRE-2 and SIE motifs. While the CRE-2-binding complex contains the CREB protein and is activated by IL-3 through the phosphatidylinositol 3-kinase/Akt-dependent pathway, the identity and cytokine activation pathway of the SIE-binding complex remains unclear. In this report, we demonstrated that PU.1 is one component of the SIE-binding complex. A chromatin immunoprecipitation assay further confirmed that PU.1 binds to the mcl-1 promoter region containing the SIE motif in vivo. While IL-3 stimulation does not significantly alter the SIE-binding activity of PU.1, it markedly increases PU.1's transactivation activity. The latter effect coincides with the increased phosphorylation of PU.1 following IL-3 activation of a p38 mitogen-activated protein kinase (p38(MAPK))-dependent pathway. A serine-to-alanine substitution at position 142 significantly weakens PU.1's ability to be phosphorylated by the p38(MAPK) immunocomplex. Furthermore, this S142A mutant is impaired in the ability to be further stimulated by IL-3 to transactivate the mcl-1 reporter through the SIE motif. Taken together, our results demonstrate that IL-3 stimulation of mcl-1 gene transcription through the SIE motif involves phosphorylation of PU.1 at serine 142 by a p38(MAPK)-dependent pathway.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Androstadienes / pharmacology
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Animals
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Cell Line / drug effects
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Cell Line / metabolism
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Cyclic AMP Response Element-Binding Protein / physiology
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Enzyme Inhibitors / pharmacology
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Genes, Immediate-Early / drug effects
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Hematopoietic Stem Cells / drug effects
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Hematopoietic Stem Cells / metabolism
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Humans
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Imidazoles / pharmacology
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Interleukin-3 / pharmacology*
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MAP Kinase Signaling System*
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Mice
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Mitogen-Activated Protein Kinases / antagonists & inhibitors
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Mitogen-Activated Protein Kinases / genetics
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Mitogen-Activated Protein Kinases / physiology*
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Mutagenesis, Site-Directed
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Myeloid Cell Leukemia Sequence 1 Protein
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Neoplasm Proteins / biosynthesis
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Neoplasm Proteins / genetics*
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Phosphatidylinositol 3-Kinases / physiology
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Phosphoinositide-3 Kinase Inhibitors
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Phosphorylation
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Promoter Regions, Genetic / drug effects*
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Protein Processing, Post-Translational
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Protein Serine-Threonine Kinases*
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Proto-Oncogene Proteins / chemistry
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Proto-Oncogene Proteins / genetics
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Proto-Oncogene Proteins / physiology*
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Proto-Oncogene Proteins c-akt
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Proto-Oncogene Proteins c-bcl-2*
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Pyridines / pharmacology
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Recombinant Fusion Proteins / physiology
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Regulatory Sequences, Nucleic Acid
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Trans-Activators / chemistry
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Trans-Activators / genetics
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Trans-Activators / physiology*
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Transcription, Genetic / drug effects*
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Wortmannin
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p38 Mitogen-Activated Protein Kinases
Substances
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Androstadienes
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Cyclic AMP Response Element-Binding Protein
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Enzyme Inhibitors
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Imidazoles
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Interleukin-3
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Mcl1 protein, mouse
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Myeloid Cell Leukemia Sequence 1 Protein
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Neoplasm Proteins
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Phosphoinositide-3 Kinase Inhibitors
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Proto-Oncogene Proteins
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Proto-Oncogene Proteins c-bcl-2
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Pyridines
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Recombinant Fusion Proteins
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Trans-Activators
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proto-oncogene protein Spi-1
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AKT1 protein, human
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Protein Serine-Threonine Kinases
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Proto-Oncogene Proteins c-akt
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Mitogen-Activated Protein Kinases
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p38 Mitogen-Activated Protein Kinases
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SB 203580
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Wortmannin