Analysis of the mechanism for chromatin remodeling in embryos reconstructed by somatic nuclear transfer.
The objective of the present study was to understand the molecular/biochemical nature of chromatin remodeling that occurs in the somatic nuclei transferred into oocytes. We produced the reconstructed mouse embryos by two different protocols of nuclear transfer. The nucleus of a cumulus cell was transferred into enucleated unfertilized oocytes (transferred before activation, TA protocol) or activated oocytes (activated before transfer, AT protocol). More than half (56.1%) of the embryos reconstructed using the TA protocol developed to the morula/blastocyst stage, whereas very few (1.0%) of the embryos reconstructed using the AT protocol reached the morula/blastocyst stage. These embryos were analyzed for the events associated with transcriptional regulation. Changes in transcriptional activity, nuclear accumulation of TATA box binding protein (TBP), and DNase I sensitivity were examined after nuclear transfer. In the embryos reconstructed by TA protocol, all of these events occurred in a manner similar to that in the control diploid parthenogenetic embryos. The transcriptional activity was silenced after nuclear transfer and resumed at the late 1-cell stage. TBP was displaced and subsequently accumulated at the early and the late 1-cell stage, respectively. DNase I sensitivity was increased and then decreased at the early and late 1-cell stage, respectively. In contrast, embryos reconstructed using the AT protocol did not show such changes in transcriptional activity, TBP accumulation, and DNase I sensitivity. These events would be necessary for differentiated nuclei to restore totipotency and are useful indices to evaluate successful chromatin remodeling.
Department of Veterinary Science, National Institute of Infectious Diseases, Tokyo 162-8640, Japan.
Jin-Moon
JM
Kim
Atsuo
A
Ogura
Masao
M
Nagata
Fugaku
F
Aoki
D000818:Animals; D001755:Blastocyst; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000648:ultrastructure*; D019976:Cloning, Organism; D003571:Cytochalasin B / Q000494:pharmacology; D003850:Deoxyribonuclease I / Q000494:pharmacology; D004622:Embryo, Mammalian / Q000648:ultrastructure*; D005314:Embryonic and Fetal Development; D005260:Female; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D009028:Morula; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000187:drug effects / Q000648:ultrastructure*; D035181:TATA-Box Binding Protein / Q000378:metabolism
D002843:Chromatin; D035181:TATA-Box Binding Protein; D003571:Cytochalasin B; D003850:Deoxyribonuclease I