Conclusions
Fam70A regulates meiosis and quality of mouse oocytes through both canonical and non-canonical Wnt5a signalling pathways.
Methods
Co-immunoprecipitation, immunogold labelling-electron microscopy, co-localization and yeast two-hybrid assays were used to verify the interaction. Antibody or small interfering RNA transfection was used to deplete the proteins. Immunofluorescence, immunohistochemistry and live tracker staining were used to examine the localization or characterize phenotypes. Western blot was used to examine the protein level.
Results
Fam70A was enriched in oocyte membranes important for normal meiosis. Fam70A depletion remarkably disrupted spindle assembly, chromosome congression and first polar body extrusion, which subsequently increased aneuploidy and abnormal fertilization. Moreover, Fam70A directly bound Wnt5a, the most abundant Wnt member within oocytes. Depletion of either Fam70A or Wnt5a remarkably increased adenomatous polyposis coli (APC), which stabilizes active β-catenin and microtubules. Consequently, depletion of either Fam70A or Wnt5a remarkably increased p-β-catenin (inactive form) and acetylated tubulin, while APC knockdown remarkably decreased these two. Furthermore, Fam70A depletion remarkably reduced Akt phosphorylation. Conclusions: Fam70A regulates meiosis and quality of mouse oocytes through both canonical and non-canonical Wnt5a signalling pathways.