Aberrant profile of gene expression in cloned mouse embryos derived from donor cumulus nuclei

Abstract

Somatic cell nuclear transfer has successfully been used to clone several mammalian species including the mouse, albeit with extremely low efficiency. This study investigated gene expression in cloned mouse embryos derived from cumulus cell donor nuclei, in comparison with in vivo fertilized mouse embryos, at progressive developmental stages. Enucleation was carried out by the conventional puncture method rather than by the piezo-actuated technique, whereas nuclear transfer was achieved by direct cumulus nuclear injection. Embryonic development was monitored from chemically induced activation on day 0 until the blastocyst stage on day 4. Poor developmental competence of cloned embryos was observed, which was confirmed by lower cell counts in cloned blastocysts, compared with the in vivo fertilized controls. Subsequently, real-time polymerase chain reaction was used to analyze and compare embryonic gene expression at the 2-cell, 4-cell, and blastocyst stages, between the experimental and control groups. The results showed reduced expression of the candidate genes in cloned 2-cell stage embryos, as manifested by poor developmental competence, compared with expression in the in vivo fertilized controls. Cloned 4-cell embryos and blastocysts, which had overcome the developmental block at the 2-cell stage, also showed up-regulated and down-regulated expression of several genes, strongly suggesting incomplete nuclear reprogramming. We have therefore demonstrated that aberrant embryonic gene expression is associated with low developmental competence of cloned mouse embryos. To improve the efficiency of somatic cell nuclear transfer, strategies to rectify aberrant gene expression in cloned embryos should be investigated.

Differential gene expression patterns in porcine nuclear transfer embryos reconstructed with fetal fibroblasts and mesenchymal stem cells

“…This finding supports a relationship between incidence of cell death and developmental potential as indicated by the lower developmental rate in NT embryos derived from FF. Aberrant gene expression is frequently observed in NT embryos of mouse, bovine, and porcine, and is one of the suggested causes of the low success rates of this approach (Boiani et al, 2002;Bortvin et al, 2003;Zhu et al, 2004;Miyazaki et al, 2005;Li et al, 2005;Tong et al, 2006). Examination of the gene expression pattern in NT embryos would aid in determining the causes of such abnormal development.…”

“…A favored hypothesis for the developmental incompetence of clones is inadequate reprogramming of the transplanted nucleus to a state equivalent to that of an early embryonic nucleus. Nuclear reprogramming of donor cells (within the ooplasm) is a complex process involving the orderly switching off and on of a diverse array of somatic and embryonic genes, to support embryogenesis and concomitant developmental events (Tong et al, 2006). It is likely that inappropriate expression of key developmental genes may contribute to the lethality of cloned embryos (Li, 2002;Jaenisch et al, 2005).…”

Identification of Inappropriately Reprogrammed Genes by Large-Scale Transcriptome Analysis of Individual Cloned Mouse Blastocysts

“…The complete reprogramming required for normal development is induced in only a few cases; consequently, faulty epigenetic changes accompanied by diverse abnormalities in the development of somatic/embryonic stem cell nuclear transferred (SECNT) embryos occur very frequently [13], [14], [15], [16], [17], [18]. Research on pre- and postimplantation development of SECNT embryos has shown that the embryos rapidly lose their developmental ability around the time of implantation, resulting in failure of implantation and normal embryogenesis [19], [20].…”

Inheritance of Histone H3 Methylation in Reprogramming of Somatic Nuclei Following Nuclear Transfer

“…This indicates that histone methylation may be involved in embryonic gene expression. Tong et al reported that several genes required for embryonic development are aberrantly expressed in cloned mouse embryos [18]. However, it is unclear whether aberrant gene expression in cloned embryos is directly related to epigenetic reprogramming.…”