Abstract
Bone morphogenetic protein-15 (BMP-15), an oocyte growth factor belonging to the transforming growth factor-beta superfamily, has recently been shown to be necessary for normal female fertility in mammals. We have previously demonstrated that BMP-15 regulates granulosa cell (GC) proliferation and differentiation; namely, BMP-15 promotes GC mitosis, suppresses follicle-stimulating hormone (FSH) receptor expression, and stimulates kit ligand expression. Although the role of BMP-15 in female reproduction has progressively deserved much attention, there is nothing known to date about the signaling pathway and receptors for BMP-15. Using rat primary GCs and a human GC cell line, COV434, we have now found that administration of BMP-15 causes a rapid and transient phosphorylation, thus activation, of the Smad1/5/8 pathway. BMP-15 also stimulated promoter activity of a selective BMP-responsive reporter construct, further demonstrating the stimulation of Smad1/5/8 signaling by BMP-15. In contrast, BMP-15 stimulation of Smad2 phosphorylation was very weak. To identify the receptors for BMP-15, we utilized recombinant extracellular domains of individual transforming growth factor-beta superfamily receptors and found that activin receptor-like kinase-6 extracellular domain most effectively co-immunoprecipitates with BMP-15, whereas BMP receptor type II extracellular domain was most effective in inhibiting BMP-15 bioactivity on FSH-induced progesterone production and GC thymidine incorporation. We also investigated whether activation of the MAPK pathway is necessary for BMP-15 biological activity and found that the addition of U0126, an inhibitor of ERK1/2 phosphorylation, suppresses BMP-15 activity on GC mitotsis but not on FSH-induced progesterone production, suggesting a selective signaling cascade in GC proliferation and differentiation.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Activin Receptors, Type I / genetics
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Activin Receptors, Type I / metabolism
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Activins / metabolism
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Animals
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Bone Morphogenetic Protein 15
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Bone Morphogenetic Protein Receptors, Type I
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Bone Morphogenetic Protein Receptors, Type II
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Butadienes / pharmacology
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Cells, Cultured
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DNA-Binding Proteins / metabolism
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Enzyme Inhibitors / pharmacology
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Female
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Follicle Stimulating Hormone / metabolism
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Genes, Reporter
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Granulosa Cells / drug effects
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Granulosa Cells / physiology*
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Growth Differentiation Factor 9
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Humans
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Intercellular Signaling Peptides and Proteins / metabolism*
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Membrane Proteins / metabolism
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Mitogen-Activated Protein Kinases / metabolism
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Nitriles / pharmacology
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Presenilin-1
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Progesterone / metabolism
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism
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Protein Structure, Tertiary
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Rats
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Rats, Sprague-Dawley
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Receptors, Growth Factor*
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Receptors, Transforming Growth Factor beta / genetics
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Receptors, Transforming Growth Factor beta / metabolism
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Signal Transduction / physiology*
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Smad Proteins
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Smad1 Protein
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Thymidine / metabolism
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Trans-Activators / metabolism
Substances
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BMP15 protein, human
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Bmp15 protein, rat
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Bone Morphogenetic Protein 15
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Butadienes
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DNA-Binding Proteins
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Enzyme Inhibitors
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GDF9 protein, human
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Gdf9 protein, rat
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Growth Differentiation Factor 9
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Intercellular Signaling Peptides and Proteins
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Membrane Proteins
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Nitriles
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PSEN1 protein, human
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Presenilin-1
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Receptors, Growth Factor
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Receptors, Transforming Growth Factor beta
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Recombinant Fusion Proteins
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SMAD1 protein, human
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Smad Proteins
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Smad1 Protein
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Smad1 protein, rat
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Trans-Activators
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U 0126
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Activins
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Progesterone
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Follicle Stimulating Hormone
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Protein Serine-Threonine Kinases
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Mitogen-Activated Protein Kinases
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Activin Receptors, Type I
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BMPR2 protein, human
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Bmpr2 protein, rat
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Bone Morphogenetic Protein Receptors, Type I
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Bone Morphogenetic Protein Receptors, Type II
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Thymidine