Preimplantation expression of the somatic form of Dnmt1 suggests a role in the inheritance of genomic imprints

BACKGROUND: Identical DNA methylation differences between maternal and paternal alleles in gametes and adults suggest that the inheritance of genomic imprints is strictly due to the embryonic maintenance of DNA methylation. Such maintenance would occur in association with every cycle of DNA replication, including those of preimplantation embryos. RESULTS: The expression of the somatic form of the Dnmt1 cytosine methyltransferase (Dnmt1s) was examined in cleavage-stage preimplantation mouse embryos. Low concentrations of Dnmt1s are found in 1-, 2-, 4-, and 8-cell embryos, as well as in morulae and blastocysts. Dnmt1s is present in the cytoplasm at all stages, and in the nuclei of all stages except the 1-cell, pronuclear-stage embryo. The related oocyte-derived Dnmt1o protein is also present in nuclei of 8-cell embryos, along with embryo-synthesized Dnmt1s. Dnmt1s protein expressed in 1-cell and 2-cell embryos is derived from the oocyte, whereas the embryo synthesizes its own Dnmt1s from the 2-cell stage onward. CONCLUSION: These observations suggest that Dnmt1s provides maintenance methyltransferase activity for the inheritance of methylation imprints in the early mouse embryo. Moreover, the ability of Dnmt1o and Dnmt1s proteins synthesized at the same time to substitute for one another's maintenance function, but the lack of functional interchange between oocyte- and embryo-synthesized Dnmt1 proteins, suggests that the developmental source is the critical determinant of Dnmt1 function during preimplantation development

Lhx1 Is Required for Specification of the Renal Progenitor Cell Field

In the vertebrate embryo, the kidney is derived from the intermediate mesoderm. The LIM-class homeobox transcription factor lhx1 is expressed early in the intermediate mesoderm and is one of the first genes to be expressed in the nephric mesenchyme. In this study, we investigated the role of Lhx1 in specification of the kidney field by either overexpressing or depleting lhx1 in Xenopus embryos or depleting lhx1 in an explant culture system. By overexpressing a constitutively-active form of Lhx1, we established its capacity to expand the kidney field during the specification stage of kidney organogenesis. In addition, the ability of Lhx1 to expand the kidney field diminishes as kidney organogenesis transitions to the morphogenesis stage. In a complimentary set of experiments, we determined that embryos depleted of lhx1, show an almost complete loss of the kidney field. Using an explant culture system to induce kidney tissue, we confirmed that expression of genes from both proximal and distal kidney structures is affected by the absence of lhx1. Taken together our results demonstrate an essential role for Lhx1 in driving specification of the entire kidney field from the intermediate mesoderm

Loss of DNMT1o Disrupts Imprinted X Chromosome Inactivation and Accentuates Placental Defects in Females

The maintenance of key germline derived DNA methylation patterns during preimplantation development depends on stores of DNA cytosine methyltransferase-1o (DNMT1o) provided by the oocyte. Dnmt1omat-/- mouse embryos born to Dnmt1Δ1o/Δ1o female mice lack DNMT1o protein and have disrupted genomic imprinting and associated phenotypic abnormalities. Here, we describe additional female-specific morphological abnormalities and DNA hypomethylation defects outside imprinted loci, restricted to extraembryonic tissue. Compared to male offspring, the placentae of female offspring of Dnmt1Δ1o/Δ1o mothers displayed a higher incidence of genic and intergenic hypomethylation and more frequent and extreme placental dysmorphology. The majority of the affected loci were concentrated on the X chromosome and associated with aberrant biallelic expression, indicating that imprinted X-inactivation was perturbed. Hypomethylation of a key regulatory region of Xite within the X-inactivation center was present in female blastocysts shortly after the absence of methylation maintenance by DNMT1o at the 8-cell stage. The female preponderance of placental DNA hypomethylation associated with maternal DNMT1o deficiency provides evidence of additional roles beyond the maintenance of genomic imprints for DNA methylation events in the preimplantation embryo, including a role in imprinted X chromosome inactivation. © 2013 McGraw et al

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Conserved features of imprinted differentially methylated domains

Genomic imprinting is a conserved epigenetic phenomenon in eutherian mammals, with regards both to the genes that are imprinted and the mechanism underlying the expression of just one of the parental alleles. Epigenetic modifications of alleles of imprinted genes are established during oogenesis and spermatogenesis, and these modifications are then inherited. Differentially methylated domains (DMDs) of imprinted genes are the genomic sites of these inherited epigenetic imprints. We previously showed that CpG-rich imperfect tandem direct repeats within three different mouse DMDs (Snurf/Snrpn, Kcnq1 and Igf2r), each with a unique sequence, play a central role in maintaining the differential methylation. This finding implicates repeat-related DNA structure, not sequence, in the imprinting mechanism. To better define the important features of this signal, we compared sequences of these three DMD tandem repeats among mammalian species. All DMD repeats contain short indirect repeats, many of which are organized into larger unit repeats. Even though the larger repeat units undergo deletion and addition during evolution (most likely through unequal crossovers during meiosis), the size of DMD tandem repeated regions has remained remarkably stable during mammalian evolution. Moreover, all three DMD tandem repeats have a high-CpG content, an ordered arrangement of CpG dinucleotides, and similar predicted secondary structures. These observations suggest that a structural feature or features of these DMD tandem repeats is the conserved DMD imprinting signal

Preimplantation expression of the somatic form of Dnmt1 suggests a role in the inheritance of genomic imprints

Background: Identical DNA methylation differences between maternal and paternal alleles in gametes and adults suggest that the inheritance of genomic imprints is strictly due to the embryonic maintenance of DNA methylation. Such maintenance would occur in association with every cycle of DNA replication, including those of preimplantation embryos. Results: The expression of the somatic form of the Dnmt1 cytosine methyltransferase ( Dnmt1s) was examined in cleavage-stage preimplantation mouse embryos. Low concentrations of Dnmt1s are found in 1-, 2-, 4-, and 8-cell embryos, as well as in morulae and blastocysts. Dnmt1s is present in the cytoplasm at all stages, and in the nuclei of all stages except the 1-cell, pronuclear-stage embryo. The related oocyte-derived Dnmt1o protein is also present in nuclei of 8-cell embryos, along with embryo-synthesized Dnmt1s. Dnmt1s protein expressed in 1- cell and 2- cell embryos is derived from the oocyte, whereas the embryo synthesizes its own Dnmt1s from the 2- cell stage onward. Conclusion: These observations suggest that Dnmt1s provides maintenance methyltransferase activity for the inheritance of methylation imprints in the early mouse embryo. Moreover, the ability of Dnmt1o and Dnmt1s proteins synthesized at the same time to substitute for one another's maintenance function, but the lack of functional interchange between oocyte-and embryo- synthesized Dnmt1 proteins, suggests that the developmental source is the critical determinant of Dnmt1 function during preimplantation development

Preimplantation expression of the somatic form of Dnmt1 suggests a role in the inheritance of genomic imprints-3

H the UPT82 antiserum. Mo = morula. Bl = blastocyst. B. Time course of Dnmt1s expression during preimplantation development in embryos derived from crosses between wild-type (+/+) female mice and homozygous (V/V) male mice. Inset is bright-field image of immunostained 1-cell embryo. C. Time courses of Dnmt1s expression during preimplantation development in embryos derived from crosses between heterozygous (V/+) female mice and homozygous (V/V) male mice. The 4-cell and 8-cell embryos in the top row show primarily nuclear Dnmt1s staining whereas 4-cell and 8-cell embryos in the botton row show little or no nuclear Dnmt1s. All seven 2-cell, three out of seven 4-cell and six out of 13 8-cell embryos showed nuclear staining. Bar, 20 μm.<p><b>Copyright information:</b></p><p>Taken from "Preimplantation expression of the somatic form of Dnmt1 suggests a role in the inheritance of genomic imprints"</p><p>http://www.biomedcentral.com/1471-213X/8/9</p><p>BMC Developmental Biology 2008;8():9-9.</p><p>Published online 25 Jan 2008</p><p>PMCID:PMC2266903.</p><p></p

Preimplantation expression of the somatic form of Dnmt1 suggests a role in the inheritance of genomic imprints-2

UPT82 antiserum. Intracellular DNA was detected with DAPI. B. Immunostaining of MII oocytes and pronuclear-stage embryos of different genotypes with the UPT82 antiserum. PN3 and PN4 refer to pronuclear stage 3 and 4 as defined by morphologic criteria [17]. Insets are bright-field images of immunostained embryos. C. Immunostaining of pronuclear-stage embryos with the UPTC21 antiserum to detect the abundant Dnmt1o protein. Genotype abbreviations are the same as in the legends to Figures 1 and 2. Bar, 20 μm.<p><b>Copyright information:</b></p><p>Taken from "Preimplantation expression of the somatic form of Dnmt1 suggests a role in the inheritance of genomic imprints"</p><p>http://www.biomedcentral.com/1471-213X/8/9</p><p>BMC Developmental Biology 2008;8():9-9.</p><p>Published online 25 Jan 2008</p><p>PMCID:PMC2266903.</p><p></p