In our previous work, we carried out microarray-based global expression profiling of all preimplantation stages in mouse. They revealed and characterized distinctive patterns of maternal RNA degradation and two major transient waves of de novo transcription. The first wave corresponds to zygotic genome activation (ZGA);the second wave, named mid-preimplantation gene activation (MGA), precedes the dynamic morphological and functional changes from the morula to blastocyst stage. We also developed a technique to do large-scale Whole Mount In Situ Hybridization (WISH), which allows us to reveal the spatial expression patterns of 91 genes in mouse preimplantation embryos. Through these analyses, we identified a number of genes that show unique expression patterns. As an example of ZGA genes, we found Zscan4, which is expressed exclusively in 2-cell mouse embryos and ES cells and plays a critical role in the progression of preimplantation development. As an example of MGA genes, we found Trim43. We showed that the downregulation of Zscan4 expression in 2-cell embryos by shRNAs delays the normal transition from 2-cell embryos to 4-cell embryos for about 24 hours, resulting in abnormal blastocysts that failed to implant in the uterus. This indicates a critical role of Zscan4 in preimplantation development. We also found that Zscan4 is expressed only in late 2-cell embryos and in the subpopulation (1 - 5%) of ES cells maintained in the undifferentiated condition. We found that other 2-cell-specific genes are also highly expressed in Zscan4(+) cells. This indicates that the subpopulation of ES cells marked by Zscan4 expression shows some resemblance to 2-cell embryos, whereas the majority of ES cells show similarities to the Inner Cell Mass (ICM) cells in blastocysts. Zscan4 is thus associated with a unique transient state in undifferentiated ES cells, in which other 2-cell embryo-specific genes are activated. By carrying out cell lineage tracing experiments, we have found that, although Zscan4 is expressed only in 5% of ES cells in culture at a given time, essentially all ES cells experience a Zscan4-positive state by 9 passages. By knocking down Zscan4 activation with an shRNA, we have found that Zscan4 is essential for the long-term maintenance of genomic integrity and for mediating a regulated telomere recombination in normal undifferentiated ES cells, therefore making it indispensable for the proper long-term self-renewal in ES cells. We have also found that Zscan4 is involved in the induction and recruitment of the meiosis-specific homologous recombination machinery to telomeres. Recently we showed that Zscan4 can be used as one of the reprogramming factors and enhances the quantity and quality of induced pluripotent stem (iPS) cells. We found that nascent protein synthesis is globally repressed in the Zscan4(+) ES cells, mediated by the transient expression of newly identified eukaryotic translation initiation factor 1A (Eif1a)-like genes. Eif1a-like genes, clustered on Chromosome 12, show high sequence similarity to Eifa1 and comprise 10 genes (Eif1al1-Eif1al10) and 9 pseudogenes (Eif1al-ps1-Eif1al-ps9). We have shown that Eif1a-like genes are expressed mostly in two-cell stage mouse embryos and in the Zscan4+ state of ES cells. We also found that increasing the frequency of Zscan4 activation in mouse ES cells restores and maintains their developmental potency in long-term cell culture. We found that the potency of ES cells increased to such a high level that the injection of even a single ES cell into a tetraploid blastocyst gives rise to an entire embryo with a higher success rate.

National Institute of Health (NIH)
National Institute on Aging (NIA)
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
Application #
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
National Institute on Aging
Zip Code
Amano, Tomokazu; Hirata, Tetsuya; Falco, Geppino et al. (2013) Zscan4 restores the developmental potency of embryonic stem cells. Nat Commun 4:1966
Ko, Shigeru B H; Azuma, Sakiko; Yokoyama, Yukihiro et al. (2013) Inflammation increases cells expressing ZSCAN4 and progenitor cell markers in the adult pancreas. Am J Physiol Gastrointest Liver Physiol 304:G1103-16
Hung, Sandy S C; Wong, Raymond C B; Sharov, Alexei A et al. (2013) Repression of global protein synthesis by eif1a-like genes that are expressed specifically in the two-cell embryos and the transient zscan4-positive state of embryonic stem cells. DNA Res 20:391-402
Zalzman, Michal; Falco, Geppino; Sharova, Lioudmila V et al. (2010) Zscan4 regulates telomere elongation and genomic stability in ES cells. Nature 464:858-63
Stanghellini, Ilaria; Falco, Geppino; Lee, Sung-Lim et al. (2009) Trim43a, Trim43b, and Trim43c: Novel mouse genes expressed specifically in mouse preimplantation embryos. Gene Expr Patterns 9:595-602