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 1 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 1 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 1 sensitivity. These events would be necessary for differentiated nuclei to restore totipotency and are useful indices to evaluate successful chromatin remodeling.