title: Selection of the dominant follicle and insulin-like growth factor (IGF)-binding proteins: evidence that pregnancy-associated plasma protein A contributes to proteolysis of IGF-binding protein 5 in bovine follicular fluid.
abstract: Development of a dominant follicle is associated with decreased intrafollicular low molecular weight IGF-binding proteins (namely IGFBP-2, -4, and -5) and increased proteolysis of IGFBP-4 by pregnancy-associated plasma protein A (PAPP-A). In addition to IGFBP-4 proteolytic activity, bovine follicular fluid contains strong proteolytic activity for IGFBP-5, but not for IGFBP-2. Here we show that the IGFBP-5 protease present in bovine follicular fluid is a neutral/basic pH-favoring, Zn(2+) metalloprotease very similar to the previously described IGFBP-4 protease. We hypothesized that immunoneutralization and immunoprecipitation with anti-PAPP-A antibodies would result in abrogation of the IGFBP-4, but not the IGFBP-5, proteolytic activity in follicular fluid. As expected, anti-PAPP-A antibodies were able to neutralize and precipitate the IGFBP-4, but not the IGFBP-5, proteolytic activity of human pregnancy serum, which was used as a positive control for PAPP-A. Surprisingly, immunoneutralization and immunoprecipitation of follicular fluid from bovine preovulatory follicles with anti-PAPP-A antibodies abrogated both IGFBP-4 and IGFBP-5 proteolysis. Quantitative results derived from phosphorimaging revealed a complete inhibition of both IGFBP-4 and -5 proteolysis by follicular fluid incubated for 2 or 5 h in the presence of anti-PAPP-A antibodies. After 18 h of incubation, anti-PAPP-A antibodies still inhibited IGFBP-5 degradation, although with an efficiency lower than that for IGFBP-4 degradation. Both proteolytic activities have identical electrophoretic mobility, and a single band ( approximately 400 kDa) was detected by Western immunoblotting of bovine follicular fluid with anti-PAPP-A antibodies. Proteolysis of IGFBP-5 was readily detectable in follicular fluid from dominant follicles and was negligible in subordinate follicles from the same cohort. These results suggest that an active intrafollicular IGFBP-4/-5 proteolytic system, in which PAPP-A is the major protease involved, is an important determinant of follicular fate.
authors.affiliation: Department of Biomedical Sciences, Cornell University, Ithaca, New York 14853, USA.
authors.forename: G M
authors.initials: GM
authors.lastname: Rivera
authors.forename: J E
authors.initials: JE
authors.lastname: Fortune
mesh_terms: D000818:Animals; D000918:Antibody Specificity; D002417:Cattle; D003338:Corpus Luteum / Q000502:physiology; D004586:Electrophoresis; D005260:Female; D015571:Follicular Fluid / Q000000981:diagnostic imaging / Q000378:metabolism*; D005498:Follicular Phase; D006801:Humans; D018971:Insulin-Like Growth Factor Binding Protein 2 / Q000378:metabolism; D018974:Insulin-Like Growth Factor Binding Protein 4 / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D008970:Molecular Weight; D006080:Ovarian Follicle / Q000000981:diagnostic imaging / Q000378:metabolism*; D011233:Precipitin Tests; D011247:Pregnancy; D011266:Pregnancy-Associated Plasma Protein-A / Q000276:immunology / Q000378:metabolism*; D014463:Ultrasonography
chemical_list: D018971:Insulin-Like Growth Factor Binding Protein 2; D018974:Insulin-Like Growth Factor Binding Protein 4; D018975:Insulin-Like Growth Factor Binding Protein 5; D011266:Pregnancy-Associated Plasma Protein-A