The bioactivity of human bone morphogenetic protein-15 is sensitive to C-terminal modification: characterization of the purified untagged processed mature region.
Mol Cell Endocrinol
; 332(1-2): 106-15, 2011 Jan 30.
Article
in En
| MEDLINE
| ID: mdl-20937357
ABSTRACT
Oocyte-derived bone morphogenetic protein-15 (BMP15) is critical for the regulation of mammalian fertility. Previously we have found that a C-terminal His(6)-tag destroys the bioactivity of growth differentiation-9 (GDF9, a homolog of BMP15). In this study we found that recombinant human BMP15 is produced by HEK-293T cells in an active form, but the bioactivity is lost by C-terminal modification, specifically, fusion to a Flag tag. After purification the mature BMP15 wt is active in transcriptional reporter assays specific for Smad1/5/8 in human granulosa-luteal (hGL) and COV434 granulosa tumor cells, whereas BMP15 with a carboxy-terminal Flag tag remains inactive. Using these same cell models we found that treatment with purified mature BMP15 wt causes a rapid phosphorylation of Smad1. The purified BMP15 wt is a potent stimulator of rat granulosa cell DNA synthesis, which could be antagonized by the BMPRII ectodomain-Fc fusion molecule, whereas the BMP15C-Flag was completely inactive. Further, the BMP15 wt form is a potent stimulator of inhibin B production in hGL cells. We found that the purified BMP15 wt consists of P16 and -17, both of which are post-translationally modified forms. This is the first characterization of a purified untagged human BMP15 mature region, which is stable and highly bioactive in human and rodent granulosa cells and as such is of importance for studies on human fertility.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Bone Morphogenetic Protein 15
Type of study:
Diagnostic_studies
/
Prognostic_studies
Limits:
Animals
/
Female
/
Humans
Language:
En
Journal:
Mol Cell Endocrinol
Year:
2011
Document type:
Article
Affiliation country: