title: Signalling pathways involved in the cooperative effects of ovine and murine GDF9+BMP15-stimulated thymidine uptake by rat granulosa cells.
abstract: Growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) are oocyte-secreted factors known to be involved in regulating the proliferation and differentiation of granulosa cells during follicular growth. The aims of this study were to determine the signalling pathways used by recombinant forms of murine and ovine GDF9 and BMP15 in combination (GDF9+BMP15) and the molecular complexes formed by combinations of these factors. Differences in the molecular forms of combinations of murine and ovine GDF9+BMP15 were observed by western blot analysis. Ovine GDF9+BMP15-stimulated (3)H-thymidine uptake was completely blocked by SMAD2/3 and nuclear factor-κB pathway inhibitors and partially blocked by a p38-mitogen-activated protein kinase (MAPK) inhibitor. Thymidine uptake by murine GDF9+BMP15 was reduced by the SMAD2/3 and extracellular signal-regulated kinase-MAPK pathway inhibitors and increased after addition of a c-Jun N-terminal kinase inhibitor. Stimulation of (3)H-thymidine uptake by GDF9+BMP15 from either species was not affected by the SMAD1/5/8 pathway inhibitor. In conclusion, both murine and ovine GDF9+BMP15-stimulated thymidine incorporation in rat granulosa cells was dependent on the SMAD2/3 signalling pathway but not the SMAD1/5/8 pathway. Divergence in the non-SMAD signalling pathways used by murine and ovine GDF9+BMP15 was also evident and may be due to the differences observed in the molecular complexes formed by these factors. These results are consistent with the hypothesis that the disparate cooperative functions of GDF9 and BMP15 in different species are mediated by divergent non-SMAD signalling pathways.
authors.affiliation: AgResearch, Reproductive Biology, Invermay Agricultural Centre, Mosgiel, New Zealand.
authors.forename: Karen L
authors.initials: KL
authors.lastname: Reader
authors.forename: Derek A
authors.initials: DA
authors.lastname: Heath
authors.forename: Stan
authors.initials: S
authors.lastname: Lun
authors.forename: C Joy
authors.initials: CJ
authors.lastname: McIntosh
authors.forename: Andrea H
authors.initials: AH
authors.lastname: Western
authors.forename: Roger P
authors.initials: RP
authors.lastname: Littlejohn
authors.forename: Kenneth P
authors.initials: KP
authors.lastname: McNatty
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Juengel
mesh_terms: D000818:Animals; D055430:Bone Morphogenetic Protein 15 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D017077:Culture Media, Conditioned / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D005260:Female; D006107:Granulosa Cells / Q000187:drug effects / Q000378:metabolism*; D055429:Growth Differentiation Factor 9 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D034561:Sheep, Domestic; D015398:Signal Transduction / Q000187:drug effects; D051899:Smad2 Protein / Q000037:antagonists & inhibitors / Q000378:metabolism; D051900:Smad3 Protein / Q000037:antagonists & inhibitors / Q000378:metabolism; D013045:Species Specificity; D013936:Thymidine / Q000378:metabolism*
chemical_list: C525902:Bmp15 protein, mouse; D055430:Bone Morphogenetic Protein 15; D017077:Culture Media, Conditioned; C525898:Gdf9 protein, mouse; D055429:Growth Differentiation Factor 9; D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins; D051899:Smad2 Protein; C497784:Smad2 protein, rat; D051900:Smad3 Protein; C497787:Smad3 protein, rat; D004247:DNA; D013936:Thymidine