A deficiency in the mechanism for p34cdc2 protein kinase activation in mouse embryos arrested at 2-cell stage.
Mouse embryos of the ddY strain fertilized in vitro undergo the first cleavage to the 2-cell stage but not the second cleavage even 45 hr after insemination (2-cell block). We examined the phosphorylation state of p34cdc2 and histone H1 kinase activity in mouse 2-cell embryos to investigate the relationship of p34cdc2 with 2-cell block. In the first mitotic cell cycle, the amount of phosphorylated forms of p34cdc2, which were detected as the bands of retarded mobility on SDS-PAGE followed by immunoblotting with anti-p34cdc2 antibody, increased during interphase and abruptly decreased at M phase. Concomitant with this dephosphorylation, histone H1 kinase activity was increased. After the embryos cleaved to the 2-cell stage, the amounts of phosphorylated forms of p34cdc2 increased up to 33 hr after insemination. However, the activation of histone H1 kinase did not occur and the states of phosphorylation of p34cdc2 did not show any significant changes until 45 hr. In contrast, 2-cell embryos of B6C3F1 mice, which do not show a 2-cell block and develop normally to blastocysts in vitro, exhibit the dephosphorylation of p34cdc2 and an increase in histone H1 kinase activity between 31 and 45 hr after insemination. When the ddY mouse embryos arrested at the 2-cell stage were treated with okadaic acid, an inhibitor of protein phosphatases 1 and 2A, the dephosphorylation of p34cdc2 occurred and histone H1 kinase activity increased. The chromosomes of these embryos stained with 4',6'-diamidino-2-phenylindole revealed the initiation of condensation. These results suggest that 2-cell-blocked embryos contain enough p34cdc2 to induce mitotic events but the protein remains in a latent form.
Aoki
F
F
Department of Animal Breeding, Faculty of Agriculture, University of Tokyo, Japan.
Choi
T
T
Mori
M
M
Yamashita
M
M
Nagahama
Y
Y
Kohmoto
K
K
D000818:Animals; D001755:Blastocyst / Q000378:metabolism*; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D004988:Ethers, Cyclic; D016200:Maturation-Promoting Factor / Q000378:metabolism*; D051379:Mice; D008815:Mice, Inbred Strains; D008938:Mitosis; D019319:Okadaic Acid; D010766:Phosphorylation
D004988:Ethers, Cyclic; D019319:Okadaic Acid; D016203:CDC2 Protein Kinase; D016200:Maturation-Promoting Factor