Figure 1: ZP3 is concentrated in the nucleus before meiosis then relocates to the zona pellucida during meiosis.

(A) ZP3 Immunofluorescene in intact mouse oocytes showed that Gm364 concentrates within nucleus region before GVBD and levels up within zona pellucida (ZP) after GVBD in mouse oocytes. (B) Ratio of ZP3 fluorescence intensity of ZP/cytoplasm at different meiotic stages showed that ZP3 abundance within ZP was very low at germinal vesicle (GV) stage and significantly leveled up after GVBD. (C) Ratio of ZP3 fluorescence intensity of chroma/cytoplasm at different meiotic stages showed that ZP3 abundance within chromosome region was very high at germinal vesicle (GV) stage and significantly decreased after GVBD. “Chroma” here is a broad definition of chromatin region (GV stage) or regions of chomosome with surrounding spindle microtubules (GVBD, MI, MII). (D) In ZP-free GV oocytes, ZP3 still highly concentrated within nucleus region. (E,F) Western blot showed that total ZP3 in ZP-intact oocytes remained constant, while in ZP-free oocytes, ZP3 amount within cytoplasm was the highest at GV stages, then dramatically decreased after GVBD. In (E), two images of the same ZP3 blot at different exposure time, 15 sec and 60 sec were shown. (G,H) At GV stage, ZP3 localization within nucleus was distinct from Nup93 or lamin A/C. DNA in blue, ZP3 in green, Nup93 or Lamin A/C in red. Different letters in (B,C) indicate significant difference. Scale bar, 20 μm.