title: BMP signalling in human fetal ovary somatic cells is modulated in a gene-specific fashion by GREM1 and GREM2.
abstract: Do changes in the expression of bone morphogenetic proteins (BMPs) 2 and 4, and their antagonists Gremlin1 (GREM1) and Gremlin2 (GREM2) during human fetal ovarian development impact on BMP pathway activity and lead to changes in gene expression that may influence the fate and/or function of ovarian somatic cells?
  
  
  
  BMPs 2 and 4 differentially regulate gene expression in cultured human fetal ovarian somatic cells. Expression of some, but not all BMP target genes is antagonised by GREM1 and GREM2, indicating the existence of a mechanism to fine-tune BMP signal intensity in the ovary. Leucine-rich repeat-containing G-protein coupled receptor 5 (LGR5), a marker of immature ovarian somatic cells, is identified as a novel transcriptional target of BMP4.
  
  
  
  Extensive re-organisation of the germ and somatic cell populations in the feto-neonatal ovary culminates in the formation of primordial follicles, which provide the basis for a female's future fertility. BMP growth factors play important roles at many stages of ovarian development and function. GREM1, an extracellular antagonist of BMP signalling, regulates the timing of primordial follicle formation in the mouse ovary, and mRNA levels of BMP4 decrease while those of BMP2 increase prior to follicle formation in the human fetal ovary.
  
  
  
  Expression of genes encoding BMP pathway components, BMP antagonists and markers of ovarian somatic cells were determined by quantitative (q)RT-PCR in human fetal ovaries (from 8 to 21 weeks gestation) and fetal ovary-derived somatic cell cultures. Ovarian expression of GREM1 protein was confirmed by immunoblotting. Primary human fetal ovarian somatic cell cultures were derived from disaggregated ovaries by differential adhesion and cultured in the presence of recombinant human BMP2 or BMP4, with or without the addition of GREM1 or GREM2.
  
  
  
  We demonstrate that the expression of BMP antagonists GREM1, GREM2 and CHRD  increases in the lead-up to primordial follicle formation in the human fetal ovary, and that the BMP pathway is active in cultured ovarian somatic cells. This leads to differential changes in the expression of a number of genes, some of which are further modulated by GREM1 and/or GREM2. The positive transcriptional regulation of LGR5 (a marker of less differentiated somatic cells) by BMP4 in vitro suggests that increasing levels of GREM1 and reduced levels of BMP4 as the ovary develops in vivo may act to reduce LGR5 levels and allow pre-granulosa cell differentiation.
  
  
  
  While we have demonstrated that markers of different somatic cell types are expressed in the cultured ovarian somatic cells, their proportions may not represent the same cells in the intact ovary which also contains germ cells.
  
  
  
  This study extends previous work identifying germ cells as targets of ovarian BMP signalling, and suggests BMPs may regulate the development of both germ and somatic cells in the developing ovary around the time of follicle formation.
  
  
  
  Not applicable.
  
  
  
  This work was supported by The UK Medical Research Council (Grant No.: G1100357 to RAA), and Medical Research Scotland (Grant No. 345FRG to AJC). The authors have no competing interests to declare.
authors.affiliation: MRC Centre for Reproductive Health, University of Edinburgh, Queen's Medical Research Institute, Edinburgh EH16 4TJ, UK.
authors.forename: Rosemary A
authors.initials: RA
authors.lastname: Bayne
authors.affiliation: MRC Centre for Reproductive Health, University of Edinburgh, Queen's Medical Research Institute, Edinburgh EH16 4TJ, UK.
authors.forename: Douglas J
authors.initials: DJ
authors.lastname: Donnachie
authors.affiliation: MRC Centre for Reproductive Health, University of Edinburgh, Queen's Medical Research Institute, Edinburgh EH16 4TJ, UK.
authors.forename: Hazel L
authors.initials: HL
authors.lastname: Kinnell
authors.affiliation: Department of Comparative Biomedical Sciences, The Royal Veterinary College, London NW1 0TU, UK.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Childs
authors.affiliation: MRC Centre for Reproductive Health, University of Edinburgh, Queen's Medical Research Institute, Edinburgh EH16 4TJ, UK Richard.Anderson@ed.ac.uk.
authors.forename: Richard A
authors.initials: RA
authors.lastname: Anderson
mesh_terms: D055415:Bone Morphogenetic Protein 4 / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation / Q000235:genetics / Q000502:physiology; D016207:Cytokines; D005260:Female; D005786:Gene Expression Regulation / Q000235:genetics; D005854:Germ Cells / Q000166:cytology / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D009865:Oocytes / Q000166:cytology / Q000378:metabolism; D006080:Ovarian Follicle / Q000166:cytology / Q000378:metabolism; D010053:Ovary / Q000166:cytology* / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology
chemical_list: D055415:Bone Morphogenetic Protein 4; D016207:Cytokines; C486399:GREM1 protein, human; C495107:GREM2 protein, human; D036341:Intercellular Signaling Peptides and Proteins
keywords: BMP; BMP antagonist; human fetal ovary; ovarian development; ovarian somatic cell; pre-granulosa cell