Protein 14-3-3 eta is essential for normal meiotic spindle assembly in mouse eggs

The 14-3-3 proteins constitute a family of conserved, acidic proteins in a variety of organisms, regulating important intracellular events including cell cycle control, apoptosis, signal transduction and embryonic development. However, little is known about the functions of 14-3-3 in mammalian reproduction. There are seven mammalian isoforms of 14-3-3 (beta, gamma, epsilon, eta, zeta, tau/theta and sigma) encoded by different genes. We previously reported accumulation and co-localization of a specific isoform of 14-3-3, namely 14-3-3 eta, in the region of meiotic spindle apparatus in mouse eggs. To determine the role of 14-3-3 eta, we microinjected mouse oocytes with a translation-blocking morpholino oligonucleotide to knock down the expression of 14-3-3 eta mRNA. Overnight incubation of the morpholino-injected oocytes followed by in vitro maturation revealed absence or impairment of meiotic spindle assembly in eggs, as shown by immunocytochemical staining of 14-3-3 eta and alpha-tubulin along with observation of chromosomes. Duolink In Situ Proximity Ligation Assay indicated marked interaction of 14-3-3 eta with alpha-tubulin in the region of meiotic spindle in all eggs examined, with prominent cortical accumulation about the spindles. These results suggest that 14-3-3 eta is necessary for normal meiotic spindle formation in mouse eggs. The study will help to elucidate the functional importance of 14-3-3 proteins in regulating mammalian oocyte maturation and female reproductive development.