Abstract
Trophoblast differentiation during placentation involves an epithelial-mesenchymal transition (EMT) with loss of E-cadherin and gain of trophoblast invasiveness. Mice harboring a point mutation that renders inactive the mitogen-activated protein kinase kinase kinase MEKK4 exhibit dysregulated placental development with increased trophoblast invasion. Isolated MEKK4 kinase-inactive trophoblast stem (TS) cells cultured under undifferentiating, self-renewing conditions in the presence of fibroblast growth factor 4 (FGF4) display increased expression of Slug, Twist, and matrix metalloproteinase 2 (MMP2), loss of E-cadherin, and hyperinvasion of extracellular matrix, each a hallmark of EMT. MEKK4 kinase-inactive TS cells show a preferential differentiation to Tpbp alpha- and Gcm1-positive trophoblasts, which are indicative of spongiotrophoblast and syncytiotrophoblast differentiation, respectively. FGF4-stimulated Jun N-terminal kinase (JNK) and p38 activity is markedly reduced in MEKK4 kinase-inactive TS cells. Chemical inhibition of JNK in wild-type TS cells induced a similar EMT response as loss of MEKK4 kinase activity, including inhibition of E-cadherin expression and increased expression of Slug, MMP2, Tpbp alpha, and Gcm1. Chromatin immunoprecipitation analyses revealed changes in AP-1 composition with increased Fra-2 and decreased Fra-1 and JunB binding to the regulatory regions of Gcm1 and MMP2 genes in MEKK4 kinase-inactive TS cells. Our results define MEKK4 as a signaling hub for FGF4 activation of JNK that is required for maintenance of TS cells in an undifferentiated state.
Publication types
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Research Support, N.I.H., Extramural
MeSH terms
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Activins / genetics
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Activins / metabolism
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Animals
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Cadherins / metabolism
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Cathepsins / genetics
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Cathepsins / metabolism
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Cell Differentiation / physiology
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Cells, Cultured
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DNA-Binding Proteins
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Embryo, Mammalian* / cytology
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Embryo, Mammalian* / physiology
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Enzyme Activation
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Extracellular Matrix
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Female
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Fibroblast Growth Factor 4 / genetics
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Fibroblast Growth Factor 4 / metabolism*
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JNK Mitogen-Activated Protein Kinases / genetics
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JNK Mitogen-Activated Protein Kinases / metabolism*
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MAP Kinase Kinase Kinase 4 / genetics
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MAP Kinase Kinase Kinase 4 / metabolism*
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Matrix Metalloproteinase 2 / genetics
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Matrix Metalloproteinase 2 / metabolism
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Mice
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Mice, Inbred C57BL
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Mice, Knockout
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Neuropeptides / genetics
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Neuropeptides / metabolism
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Placenta / cytology
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Pregnancy
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Signal Transduction / physiology
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Snail Family Transcription Factors
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Stem Cells / cytology
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Stem Cells / physiology*
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Transcription Factors / genetics
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Transcription Factors / metabolism
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Transforming Growth Factor beta / genetics
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Transforming Growth Factor beta / metabolism
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Trophoblasts / cytology*
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Trophoblasts / physiology
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Twist-Related Protein 1 / genetics
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Twist-Related Protein 1 / metabolism
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p38 Mitogen-Activated Protein Kinases / genetics
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p38 Mitogen-Activated Protein Kinases / metabolism
Substances
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Cadherins
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DNA-Binding Proteins
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Fibroblast Growth Factor 4
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Gcm1 protein, mouse
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Neuropeptides
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Snai2 protein, mouse
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Snail Family Transcription Factors
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Transcription Factors
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Transforming Growth Factor beta
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Twist-Related Protein 1
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Activins
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JNK Mitogen-Activated Protein Kinases
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p38 Mitogen-Activated Protein Kinases
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MAP Kinase Kinase Kinase 4
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Cathepsins
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cathepsin S
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Matrix Metalloproteinase 2