Introduction: Degradation of proteins, such as cyclin B1 and securin is a hallmark of resumption of meiosis in oocytes. In order to degrade these, as well as other substrate proteins, the oocyte employs the multiple enzymes of the ubiquitin-proteasome pathway E1, E2 and E3. Unlike the accumulated knowledge of E3, there is hardly any information related to E2 in this system.
Aims: Deciphering the specific ubiquitin-conjugating enzymes (E2s) that play a role in oocyte meiosis.
M&M: Mouse oocytes were either microinjected with specific morpholino antisense for the depletion of different E2 proteins, or by particular E2 cRNA for over-expression of the corresponding enzymes. Live imaging was employed for monitoring first polar body (PBI) extrusion, spindle formation and chromosome segregation in the microinjected oocytes. In addition, securin-GFP levels were examined in the different experiments.
Results: We found that knocking-down each of the three E2 proteins, UBE2C, UBE2S and UBE2D3 inhibited PBI extrusion, spoiled spindle formation and disturbed chromosomes segregation. We also learned that securin levels in these oocytes remained high. Alternatively, over expression of Ube2C or Ube2S resulted in a larger fraction of oocytes extruding PBI, an event that took place prematurely. In addition, oocytes over expressing Ube2C exhibited aberrations in cytokinesis, did not arrest at the second metaphase and extruded the second PB.
Conclusions: Our study identifies and characterizes the E2 proteins that participate in ubiquitination during meiosis pointing towards their role in PBI extrusion, spindle formation and appropriate chromosome segregation. These results may contribute to our understanding of aneuploidy and its consequent genetic malformations.
