Fig 2: GDF9 and BMP15 in human oocytes. (A) GDF9 was detected at a molecular weight of 47 kDa in a pool of 11 human immature oocytes. Using a blocking peptide against GDF9 (BG016, Ansh Labs) the band disappeared. BMP15 was detected at molecular weight of 43 kDa in a pool of 20 human immature oocytes. After including a BMP15 blocking peptide (BB028, Ansh Labs) the band disappeared, validating the specificity of the antibody. (B) GDF9 and BMP15 were detected after IVM in pools of 23 GV oocytes per lane and 28 MI/MII oocytes per lane. Note that the bands from recombinant GDF9 and BMP15 standards differed significantly from the native oocyte derived bands. The uncropped western blot membranes can be found in additional files 3 (A) and 4 (B)

Fig 3: Relative expression of seven genes from the GDF9 and BMP15 signaling pathways in cumulus cells from human oocytes before IVM (GV(Fresh), n = 24), and after IVM (GV, n = 21; and MII, n = 27). GV and MII respectively refer to germinal vesicle and metaphase II stages. (A) BMPR2, (B) ALK5, (C) ALK 6, (D) SMAD2, (E) SMAD3, (F) SMAD1, and (G) SMAD5. Error bars represent SEM. *P < 0.05; **P ≤ 0.005

Fig 4: GDF9 and BMP15 localization in human ovarian follicles. (A) GDF9 (green) and HSD17β1 (red). (A1-A4) cumulus oocyte complex, (A5-A8) Magnifications of the dotted boxes in A1-A4). (B) BMP15 (green) and HSD17β1 (red). (B1-B4) cumulus oocyte complex, (B5-B8) Magnifications of the dotted boxes in B1-B4)

Fig 5: BMP9 enhances the antiviral response of HFF-1 to HCMV infection.(A) Schematic representation of the workflow. After 2 h of serum starvation, HFF-1 were stimulated for 6 h with BMP4 (18 nM), BMP6 (18 nM), BMP9 (3 nM), BMP15 (18 nM), or Activin B (4 nM), or co-stimulated with IFNβ (5 ng/ml), followed by HCMV infection (MOI 0.5) for 16 h. Cells were fixed, nuclei were stained and cells were labeled for HCMV IE1+ cells as a readout for infection. (B) HCMV IE1+ cells normalized to total cell numbers and the untreated control (white column) in BMP/Activin stimulated samples (left panel) or with IFNβ co-stimulated samples (right panel). (C) HCMV IE1+ cells normalized to total cell numbers in cells pre-stimulated with either low (0.25 nM) or high (3 nM) concentrations of BMP9 (green symbols) or IFNβ co-stimulated with low and high concentrations of BMP9 (beige symbols). (D) HFF-1 were infected by centrifugal enhancement with HCMV WT (MOI 0.5) and supernatants of infected cells were collected in 6 h increments. 293T were co-transfected with expression plasmids for the BRE-Luciferase reporter and a Renilla luciferase normalization control. Twenty-four hours post transfection, 293T were either stimulated with supernatants from HCMV-infected cells, or supernatants from HCMV-infected cells incubated for 15 min at RT with an α-BMP9 antibody, for 16 h, followed by a dual-luciferase assay readout. Luciferase fold induction was calculated by dividing Renilla-normalized values from stimulated samples by the corresponding values from unstimulated samples. Data information: (B) Experiment was performed three independent times, one representative is shown. (C, D) Data are combined from two independent experiments. Student’s t test (unpaired, two-tailed), n.s. not significant, *P < 0.05, **P < 0.01, ***P < 0.001. Data are shown as mean ± SD. Source data are available online for this figure.