Essential role of paternal chromatin in the regulation of transcriptional activity during mouse preimplantation development.
ABSTRACT Several lines of evidence indicate that the formation of a transcriptionally repressive state during the two-cell stage in the preimplantation mouse embryo is superimposed on the activation of the embryonic genome. However, it is difficult to determine the profile of newly synthesized (nascent) RNA during this phase because large amounts of maternal RNA accumulate in maturing oocytes to support early development. Using 5-bromouridine-5'-triphosphate labeling of RNA, we have verified that nascent RNA synthesis was repressed between the two-cell and four-cell transition in normally fertilized but not in parthenogenetic embryos. Moreover, this repression was contributed by sperm (male) chromatin, which we confirmed by studying androgenetic embryos. The source of factors responsible for repressing nascent RNA production was investigated using different stages of sperm development. Fertilization with immature round spermatids resulted in a lower level of transcriptional activity than with ICSI at the two-cell stage, and this was consistent with further repression at the four-cell stage in the ICSI group. Finally, study on DNA replication and chromatin remodeling was performed using labeled histones H3 and H4 to differentiate between male and female pronuclei. The combination of male and female chromatin appeared to decrease nascent RNA production in the fertilized embryo. This study indicates that paternal chromatin is important in the regulation of transcriptional activity during mouse preimplantation development and that this capacity is acquired during spermiogenesis.
SourceAvailable from: Yi-Hui Chen[Show abstract] [Hide abstract]
ABSTRACT: Epigenetic asymmetry has been shown to be associated with the first lineage allocation event in preimplantation development, i.e., the formation of the trophectoderm (TE) and inner cell mass (ICM) lineages in the blastocyst. Because parthenogenesis causes aberrant segregation between the TE and ICM lineages, we examined several development-associated histone modifications in parthenotes, including those involved in 1) transcriptional activation (acetylated histone H3 lysine 9 (H3K9Ac) and lysine 14 (H3K14Ac), trimethylated histone H3 lysine 4 (H3K4Me3), and dimethylated histone H3 arginine 26 (H3R26Me2) and 2) transcriptional repression (trimethylated histone H3 lysine 9 (H3K9Me3) , trimethylated histone H3and lysine 27 (H3K27Me3), and mono-ubiquitinated histone H2A lysine 119 (H2AK119u1)). Here we report that in parthenotes H3R26Me2 expression decreased from the morula stage, while expression patterns and levels of H3K9Ac, H3K27Me3, and H2AK119u1 were unchanged until the blastocyst stage, whereas H3K14Ac, H3K4Me3 and H3K9Me3 showed normal patterns and levels of expressions. Relative to the decrease of H3K9Ac in the ICM and increase in the TE of parthenotes, we detected reduced expression of Tip60 acetyltransferase and HDAC1 deacetylase in the ICM and TE of parthenotes, respectively. Relative to the decrease of H3R26Me2, we also observed decreased expression of CARM1 methyltransferase and increased expression of the Wnt effector TCF7L2 and miR-181c microRNA in parthenotes. Furthermore, relative to the decrease in H3K27Me3 and H2AK119u1, we found increased phosphorylation of Akt1 and Ezh2 in parthenogenetic TE. Therefore, our findings that histone signatures are impaired in parthenotes provide a mechanistic explanation for aberrant lineage segregation and TE defects.Stem Cells and Development 10/2014; DOI:10.1089/scd.2014.0310 · 4.20 Impact Factor
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ABSTRACT: Fertilization precisely choreographs parental genomes by using gamete-derived cellular factors and activating genome regulatory programs. However, the mechanism remains elusive owing to the technical difficulties of preparing large numbers of high-quality preimplantation cells. Here, we collected >14 × 10(4) high-quality mouse metaphase II oocytes and used these to establish detailed transcriptional profiles for four early embryo stages and parthenogenetic development. By combining these profiles with other public resources, we found evidence that gene silencing appeared to be mediated in part by noncoding RNAs and that this was a prerequisite for post-fertilization development. Notably, we identified 817 genes that were differentially expressed in embryos after fertilization compared with parthenotes. The regulation of these genes was distinctly different from those expressed in parthenotes, suggesting functional specialization of particular transcription factors prior to first cell cleavage. We identified five transcription factors that were potentially necessary for developmental progression: Foxd1, Nkx2-5, Sox18, Myod1, and Runx1. Our very large-scale whole-transcriptome profile of early mouse embryos yielded a novel and valuable resource for studies in developmental biology and stem cell research. The database is available at http://dbtmee.hgc.jp.Genes & Development 12/2013; 27(24):2736-48. DOI:10.1101/gad.227926.113 · 12.64 Impact Factor
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ABSTRACT: Background Real-time quantitative reverse-transcriptase polymerase chain reaction (RT-qPCR) is the most sensitive, and valuable technique for rare mRNA detection. However, the expression profiles of reference genes under different experimental conditions, such as different mouse strains, developmental stage, and culture conditions have been poorly studied. Results mRNA stability of the actb, gapdh, sdha, ablim, ywhaz, sptbn, h2afz, tgfb1, 18 s and wrnip genes was analyzed. Using the NormFinder program, the most stable genes are as follows: h2afz for the B6D2F-1 and C57BL/6 strains; sptbn for ICR; h2afz for KOSOM and CZB cultures of B6D2F-1 and C57BL/6 strain-derived embryos; wrnip for M16 culture of B6D2F-1 and C57BL/6 strain-derived embryos; ywhaz, tgfb1, 18 s, 18 s, ywhaz, and h2afz for zygote, 2-cell, 4-cell, 8-cell, molular, and blastocyst embryonic stages cultured in KSOM medium, respectively; h2afz, wrnip, wrnip, h2afz, ywhaz, and ablim for zygote, 2-cell, 4-cell, 8-cell, molular, and blastocyst stage embryos cultured in CZB medium, respectively; 18 s, h2afz, h2afz, actb, h2afz, and wrnip for zygote, 2-cell, 4-cell, 8-cell, molular, and blastocyst stage embryos cultured in M16 medium, respectively. Conclusions These results demonstrated that candidate reference genes for normalization of target gene expression using RT-qPCR should be selected according to mouse strains, developmental stage, and culture conditions. Electronic supplementary material The online version of this article (doi:10.1186/1756-0500-7-675) contains supplementary material, which is available to authorized users.BMC Research Notes 09/2014; 7(1):675. DOI:10.1186/1756-0500-7-675