Effects of Protein Kinase Inhibitors on Gene Activation of Early Embryos in Mouse

생쥐 초기배아의 유전자 활성에 미치는 Protein Kinase Inhibitors의 영향

Transcriptional activation of the embryonic genome initiates at 2-cell stage in mouse embryo and is characterized by the synthesis of TRC which is restricted to 2-cell stage. To investigate the roles of various protein kinases on the embryonic gene activation, the effects of protein kinase inhibitors on in vitro development and protein synthetic profiles of the early mouse embryos were examinded. None of ${\alpna}-amanitin$ which is a mRNA synthetic inhibitor, H8 which is a PKA inhibitor, and H7 which is a PKC inhibitor, affected on first cleavage of mouse 1-cell embryos in vitro. However, all of these drugs inhibited the second cleavage. When the drugs were removed following treatment for 6 hours, H8 or H7 treatment showed little inhibition on subsequent development of 1-cell embryos to 2-cell stage or further. In contrast, ${\alpna}-amanitin$ irreversibly inhibited the development of 1-cell embryos to 2-cell stage following removal of the drug. Genistein, a TPK inhibitor, inhibited both the first cleavage of 1-cell embryos and the second cleavage of 2-cell embryos, suggesting that TPK activity may be important during the early cleavages. All of the above four drugs inhibited TRC synthesis as shown by the fluorographic analysis of $[^{35}S]-Met$ labeled protein profiles. When late 1-cell embryos were treated with H7 and analyzed synthetic patterns of $[^{35}S]-Met$ labeled protein, the quantitative differences of protein synthesis on SDS-PAGE appeared on 77 kD and 33 kD region at $32{\sim}38$ hours post hCG. From these studies, transcriptional activation of embryonic genome is not essenting to the mouse 1-cell embryos to develop to 2-cell stage. Hawever, TPK activity is reguisite for both the first cleavage and second cleavage. Similarly, both PKC and PKA activities are required for the second cleavage of mouse embryos, but not for the first cleavage.