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Original Article |
1 Germline Development Group, Center for Animal Transgenesis and Germ Cell Research, New Bolton Center, University of Pennsylvania, Kennett Square, PA 19348, USA
2 Laboratorio di Biologia dello Sviluppo, Dipartimento di Biologia Animale, Universitá degli Studi di Pavia, Pavia - Italy
3 University of Pennsylvania
4 Max Planck Institute for Molecular Biomedicine
* To whom correspondence should be addressed. E-mail: mboiani{at}mpi-muenster.mpg.de.
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Abstract |
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A prevailing view of cloning by somatic-cell nuclear transfer is that 'reprogramming' of gene expression occurs during the first few hours after injection of the nucleus into an oocyte, that the process is stochastic, and that the type of 'reprogramming' needed for cloning success is 'foreign' and unlikely to be readily achieved in the ooplasm. Here, we present evidence that the release of 'reprogramming' capacity is contingent on the culture environment of the clone while the contribution of aneuploidy to altered gene expression is marginal. In particular, the rate of blastocyst formation in clones and the regional distribution of mRNA for the pluripotent stem cell marker Oct4 in clonal blastocysts was highly dependent on the culture environment after cumulus cell nuclear transfer, unlike that in genetically equivalent zygotes. Epigenetic modifications of genetically identical somatic nuclei continue after the first cell division of the clones, are amenable to a degree of experimental control, and their development to the blastocyst stage and appropriate expression of Oct4 predict further outcome such as derivation of embryonic stem (ES) cells, but not fetal development. This observation indicates that development to the blastocyst stage is not equivalent to full 'reprogramming,' and lends support to the novel concept that ES cells are not the equivalent of the inner cell mass (ICM), hence the discrepancy between ES cell derivability and fetal development of clones.
Key Words. Embryo, ES cell, nuclear transfer, Oct4, reprogramming
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