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Restricting the growth of the embryo can cause adverse whole-of-life changes in an organism’s omeostasis. Such adverse long-term consequences may occur even when growth restriction occurs only during the preimplantation period. The molecular basis for these long-term effects has not been defined, although an epigenetic mechanism is suspected. Some loci seem to be more sensitive to epigenetic perturbation than others, and the agouti viable yellow allele (Avy) is the best studied example of this. It has active (hypomethylated) and inactive (hypermethylated) epialleles. This study used the Avy model to show that growth restriction of preimplantation embryos, as provided by culture of zygotes, induced persistent epigenetic changes that resulted in altered postnatal phenotype. C57BL/6 Avy/a males were mated to ovulation-induced FVB/N females, and then either zygotes were collected and cultured for 96 h and the
resulting blastocysts were transferred to pseudopregnant recipient females, blastocysts were collected from females and transferred without embryo culture, or pregnancy was allowed to proceed after mating without intervention. Culture was in a commercial human in vitro fertilization media. The proportion of pups expressing the active (hypomethylated) epiallele and yellow coat was significantly higher following zygote culture compared to embryos that were transferred without culture (P = 0.014) or natural matings (P < 0.001). There was no difference in expression of the active epiallele in pups resulting from embryo transfer (without culture) compared to natural matings. These results show for the first time that the preimplantation embryo’s growth environment can affect the postnatal expression of a defined epigenetically sensitive allele.

Maternal and zygotic Dnmt1 are necessary and sufficient for the maintenance of DNA methylation imprints during preimplantation development
Ryutaro Hirasawa, Hatsune Chiba, Masahiro Kaneda, Shoji Tajima, En Li,
Rudolf Jaenisch, and Hiroyuki Sasaki
Genes and Development 22:1607–1616 © 2008 by Cold Spring Harbor Laboratory Press ISSN 0890-9369/08; www.genesdev.org

Parental origin-specific DNA methylation regulates the monoallelic expression of the mammalian imprinted genes. The methylation marks or imprints are established in the parental germline and maintained throughout embryonic development. However, it is unclear how the methylation imprints are maintained through extensive demethylation in cleavage-stage preimplantation embryos. Previous reports suggested that DNA methyltransferase(s) other than Dnmt1 is involved in the maintenance of the imprints during cleavage. Here we demonstrate, by using conditional knockout mice, that the other known DNA methyltransferases Dnmt3a and Dnmt3b are dispensable for the maintenance of the methylation marks at most imprinted loci. We further demonstrate that a lack of both maternal and zygotic Dnmt1 results in complete demethylation of all imprinted loci examined in blastocysts. Consistent with these results we find that zygotic Dnmt1 is expressed in the preimplantation embryo. Thus, contrary to the previous reports, Dnmt1 alone is sufficient to maintain the methylation marks of the imprinted genes.

The Maternal Nucleolus Is Essential for Early Embryonic Development
in Mammals
Sugako Ogushi, Chiara Palmieri, Helena Fulka, Mitinori Saitou,
 Takashi Miyano, Josef Fulka Jr.
Science VOL 319 1 February 2008, 613-616 www.sciencemag.org

With fertilization, the paternal and maternal contributions to the zygote are not equal. The oocyte and spermatozoon are equipped with complementary arsenals of cellular structures and molecules necessary for the creation of a developmentally competent embryo. We show that the nucleolus is exclusively of maternal origin. The maternal nucleolus is not necessary for oocyte maturation; however, it is necessary for the formation of pronuclear nucleoli after fertilization or parthenogenetic activation and is essential for further embryonic development. In addition, the nucleolus in the embryo produced by somatic cell nuclear transfer originates from the oocyte, demonstrating that the maternal nucleolus supports successful embryonic development.

Paracrine factors from cumulus-enclosed oocytes ensure the successful maturation and fertilization in vitro of denuded oocytes in the cat model
Natasha M. Godard,  Budhan S. Pukazhenthi,  David E. Wildt, and Pierre Comizzoli,
Fertility and Sterility, 2008 1 doi:10.1016/j.fertnstert.2008.05.069

Objective: To better characterize cumulus-oocyte interactions during oocyte maturation and fertilization in the cat model. Design: Experimental in vitro study. Setting: Smithsonian Institution. Animal(s): Domestic shorthair cats. Intervention(s): Groups of denuded oocytes (DOs) and cumulus-oocyte complexes (COCs) were subjected to
in vitro maturation (with or without FSH and LH, with or without the gap junction disruptor 1-heptanol, in separated groups or in coculture) and inseminated in vitro (IVF; in separated groups or in coculture). Main Outcome Measure(s): Nuclear maturation, pronuclear formation, kinetics of early embryo cleavage, and blastocyst formation and quality after different in vitro conditions were compared between DOs cultured separately and DOs cocultured with COCs. Result(s): Without FSH and LH, the removal of cumulus cells prevented spontaneous meiotic resumption in DOs. With FSH and LH, groups of DOs progressed to the metaphase I stage but fully advanced to metaphase II only in coculture with intact (nondisrupted) COCs. Groups of DOs cultured separately were fertilized poorly and exhibited no blastocyst formation. In contrast, DOs cocultured with intact COCs during in vitro maturation and IVF recovered fertilizability, and approximately 35% formed blastocysts. Conclusion(s): Paracrine factors produced by cumulus-enclosed oocytes in the cat model will help to develop synthetic media for successful in vitro culture of DOs