The feasibility of producing viable cloned mammalian offspring by transplantation of adult somatic cell nuclei into oocytes provides striking evidence of the exquisite ability mammalian oocytes to reprogram nuclear function, and of the comparative plasticity of the programmed state of a variety of adult somatic cell nuclei. The advent of these cloning technologies also provides valuable new research opportunities for investigating the molecular mechanisms through which differentiative programming can be reversed in adult somatic cell nuclei, and the molecular mechanisms by which mammalian oocytes reprogram nuclear function, which is an essential part of the process of converting sperm and egg genomes into an embryonic genome. Of special interest for understanding nuclear reprogramming is to what degree the oocyte is able to erase or modify epigenetic information that is present in adult somatic cell nuclei. We hypothesize that some forms of epigenetic inheritance, such as genomic imprinting, must be maintained in order for a somatic cell nucleus to support normal development. Other forms of epigenetic inheritance, such as those associated allelic exclusion or positional identity, most likely need to be erased. The likely fate of still other forms of epigenetic modification, such as X chromosome inactivation is unclear. We also hypothesize that failure of the oocyte to elicit the necessary epigenetic changes, or the loss of essential epigenetic information in the somatic cell, will result in reduced viability of cloned embryos. To test these hypotheses, we will determine the degree to which the DNA methylation that is associated with genomic imprinting, positional identity, and allelic exclusion is altered during cloning, and whether these changes in DNA methylation are correlated with embryo viability. We also hypothesize that, because genetic differences in the ooplasm how pronuclei are modified, these same differences will affect how somatic cell nuclei are modified, and hence the viability of the clonal embryos. This will be tested by comparing the developmental capacities of cloned embryos prepared with oocytes of specific genotypes that are known to modify pronuclei differently. These studies are relevant to problems of human infertility and a variety of diseases involving inappropriate epigenetic regulation of gene function.
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