Dynamic changes in NuMA and microtubules in monkey-rabbit nuclear transfer embryos

Area of Science:

• Reproductive Biology
• Cell Biology
• Developmental Biology

Background:

• Cloning failures in monkeys are linked to nuclear mitotic apparatus (NuMA) removal and abnormal spindle formation.
• Understanding the roles of tubulin and NuMA during the initial cell cycle of cloned embryos is crucial.

Purpose of the Study:

• To investigate the transformation and assembly of tubulin and NuMA proteins in cloned monkey embryos.
• To analyze the impact of recipient oocyte activation status on donor nucleus behavior and early embryo development.

Main Methods:

• Reconstruction of cloned monkey embryos using enucleated rabbit oocytes and monkey fibroblasts.
• Observation of microtubule organization, nuclear changes (e.g., premature chromosome condensation), and spindle formation.
• Microinjection of NuMA antibody to assess its role in spindle organization.

Main Results:

• Somatic cell centrosomes were introduced, leading to microtubule organization around the donor nucleus.
• Non-activated oocytes showed high rates of premature chromosome condensation, while pre-activated oocytes rarely did.
• Approximately one-third of reconstructed embryos exhibited normal spindle formation.
• NuMA localized to pseudo-pronuclei and subsequently to mitotic spindle poles.
• NuMA antibody injection caused spindle disorganization but did not significantly affect early cleavage.

Conclusions:

• Donor monkey fibroblast NuMA contributes to normal spindle formation in enucleated rabbit oocytes.
• Recipient oocyte activation state influences donor nucleus morphology but not early embryo development outcomes.
• Spindle and chromosome alignment defects reduce cloning efficiency but spare early cleavage stages.