Molecular forms of ruminant BMP15 and GDF9 and putative interactions with receptors
Bone morphogenetic factor 15 (BMP15) and growth differentiation factor 9 (GDF9) are oocyte-secreted factors with demonstrable effects on ovarian follicular development and ovulation rate. However, the molecular forms of BMP15 and GDF9 produced by oocytes remain unclear. The aims herein, using Western blotting (WB) procedures with specific monoclonal antibodies (mabs), were to identify the molecular forms of BMP15 and GDF9 synthesised and secreted by isolated ovine (o) and bovine (b) oocytes in vitro. The mabs were known to recognise the biological forms of BMP15 or GDF9 since they had previously been shown to inhibit their bioactivities in vitro and in vivo. Using recombinant variants of oBMP15 and oGDF9, including a cysteine mutant form of oBMP15 (S356C) and a human (h) BMP15:GDF9 heterodimer (cumulin), it was established that the mabs were able to identify monomeric, dimeric, promature and higher-molecular-weight forms of BMP15 and GDF9 and cumulin (GDF9 mab only). After using non-reducing, reducing and reducing + cross-linking conditions, the major oocyte-secreted forms of o and b BMP15 and GDF9 were the cleaved and uncleaved monomeric forms of the promature proteins. There was no evidence for dimeric or heterodimeric forms of either mature BMP15 or GDF9. From in silico modelling studies using transforming growth factor beta (TGFB), activin or BMP crystal templates, and both present and previously published data, a model is proposed to illustrate how the monomeric forms of BMP15 and GDF9 may interact with their type II and type I cell-surface receptors to initiate the synergistic actions of these growth factors.
Source: The journal of Reproductive Science