Oral Presentation The Joint Annual Scientific Meetings of the Endocrine Society of Australia and the Society for Reproductive Biology 2017

BMP15 mutations associated with female reproductive disorders reduce expression, activity, or synergy with GDF9. (#98)

Liliana C Patino 1 2 , William A Stocker 3 4 , Kelly L Walton 3 , Thomas Mueller 5 , Dulama Richani 6 , David Agapiou 6 , Robert B Gilchrist 6 , Paul Laissue 2 , Craig A Harrison 3
  1. Hudson Institute of Medical Research, Clayton, VIC, Australia
  2. Universidad del Rosario, Bogota, Colombia
  3. Department of Physiology, Monash University, Clayton, VIC, Australia
  4. Department of Chemistry and Biotechnology, Swinburne University of Technology, Hawthorn, Victoria, Australia
  5. Department of Plant Physiology and Biophysics, University Wuerzburg , Wuerzburg , Germany
  6. Discipline of Obstetrics & Gynaecology, School of Women’s & Children’s Health, University of New South Wales, Sydney, NSW, Australia

Bone morphogenetic protein 15 (BMP15) is an oocyte-specific growth factor, which, together with growth differentiation factor 9 (GDF9), regulates folliculogenesis and ovulation rate. Multiple mutations in BMP15 have been identified in women with primary ovarian insufficiency (POI), supporting a pathogenic role; however, the underlying biological mechanism of many of these mutants remains unresolved. Additionally, recent evidence supports that BMP15 can heterodimerise with GDF9 to form ‘cumulin’, resulting in synergistic activation of the SMAD 2/3 signalling pathway in granulosa cells. As such, genetic mutations in BMP15 have the potential to affect either the canonical BMP15-activated SMAD 1/5 pathway, or alternatively, the cumulin-activated SMAD 2/3 pathway.  This study aimed to determine how mutations associated with ovarian disorders alter the biological activity of human BMP15, and the ability of BMP15 to synergise with GDF9. The effects of 10 BMP15 mutations on protein production, activation of granulosa cells, and synergy with GDF9 was assessed. Analysis of the expression of these variants from mammalian cells by Western blotting revealed that three mutations (L148P, F194S, and Y235C) resulted in reduced levels of mature BMP15 protein. For other mutations (R138H, A180T, and R329H), a reduction in BMP15’s ability to activate the SMAD 1/5 pathway was observed using a luciferase reporter assay. Interestingly, three BMP15 variants (R68W, F194S, and N196K) displayed a significantly reduced ability to synergize with GDF9 in SMAD 2/3 reporter assays. This study demonstrates that mutations in BMP15 associated with female reproductive pathologies reduce mature protein production, activity, or synergy with GDF9. The latter effect is perhaps most interesting given that interactions with GDF9 most likely underlie the physiology of BMP15 in the human ovary. We are currently generating recombinant forms of human BMP15, GDF9, and cumulin, which will allow us to better understand the signalling pathways activated by these oocyte-secreted factors.