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.
Kim
Jin-Moon
JM
Department of Veterinary Science, National Institute of Infectious Diseases, Tokyo 162-8640, Japan.
Ogura
Atsuo
A
Nagata
Masao
M
Aoki
Fugaku
F
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