Book ChapterDOI
X-chromosome inactivation in mouse embryonic stem cells: analysis of histone modifications and transcriptional activity using immunofluorescence and FISH.
TLDR
This chapter outlines various techniques for analyzing X-inactivation kinetics in differentiating Embryonic stem (ES) cells, particularly on changing patterns of histone modifications during X inactivation, using immunuofluorescence combined with RNA fluorescence in situ hybridization (FISH) on interphase nuclei, as well as metaphase chromosome staining combined with DNA FISH.Abstract:
Publisher Summary This chapter outlines various techniques for analyzing X-inactivation kinetics in differentiating Embryonic stem (ES) cells. The chapter focuses particularly on changing patterns of histone modifications during X inactivation, using immunuofluorescence combined with RNA fluorescence in situ hybridization (FISH) on interphase nuclei, as well as metaphase chromosome staining combined with DNA FISH. X-chromosome inactivation provides a powerful model system to investigate the different steps in facultative heterochromatin formation. During early development, one of the two X chromosomes is transcriptionally silenced in every cell of a female embryo, thereby achieving dosage compensation between males and females for X-linked gene products. The X inactivation process is dependent on the action of a unique RNA, Xist, which coats the X chromosome in cis and induces its inactivation. Once established, the inactive state of the X chromosome is highly stable in somatic cells and is normally only reversed in the female germ line. In addition, the chapter also describes a protocol for assaying late replication timing of the inactive X chromosome. Using these techniques it is possible to determine the relative order of the following events: Xist RNA coating occurs within the first 24–48 hours of differentiation; histone H3 modifications, such as hypomethylation of Lys-4, hypoacetylation of Lys- 9, and hypermethylation of Lys-9 and Lys-27 are detectable on the X chromosome in a proportion of interphase cells as soon as Xist RNA accumulates.read more
Citations
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Journal ArticleDOI
A novel role for Xist RNA in the formation of a repressive nuclear compartment into which genes are recruited when silenced
TL;DR: It is proposed that Xist RNA has multiple functions: A-repeat-independent creation of a transcriptionally silent nuclear compartment; and A- Repeat-dependent induction of gene repression, which is associated with their translocation into this silent domain.
Journal ArticleDOI
Coregulated human globin genes are frequently in spatial proximity when active
Jill M. Brown,Joanne Leach,Joyce E. Reittie,Ann Atzberger,Jane Lee-Prudhoe,William G. Wood,Douglas R. Higgs,Francisco J. Iborra,Veronica J. Buckle +8 more
TL;DR: The findings suggest that the distribution of globin genes within erythroblast nuclei is the result of a self-organizing process, involving transcriptional status, diffusional ability of chromatin, and physical interactions with nuclear proteins, rather than a directed form of higher-order control.
Journal ArticleDOI
Noncoding RNAs and Epigenetic Mechanisms During X-Chromosome Inactivation
Anne-Valerie Gendrel,Edith Heard +1 more
TL;DR: The diverse strategies used by nonplacental mammals for X-chromosome dosage compensation are described and the common features and differences between eutherians and metatherians are highlighted, in particular regarding the involvement of long noncoding RNAs.
Journal ArticleDOI
Generation of human oogonia from induced pluripotent stem cells in vitro
Chika Yamashiro,Kotaro Sasaki,Yukihiro Yabuta,Yoji Kojima,Tomonori Nakamura,Ikuhiro Okamoto,Shihori Yokobayashi,Yusuke Murase,Yukiko Ishikura,Kenjiro Shirane,Hiroyuki Sasaki,Takuya Yamamoto,Takuya Yamamoto,Mitinori Saitou +13 more
TL;DR: It is shown that hPGCLCs differentiate progressively into oogonia-like cells during a long-term in vitro culture in xenogeneic reconstituted ovaries with mouse embryonic ovarian somatic cells, establishing the germline competence of hPSCs and provide a critical step toward human in vitro gametogenesis.
Journal ArticleDOI
Smchd1-dependent and -independent pathways determine developmental dynamics of CpG island methylation on the inactive X chromosome.
Anne-Valerie Gendrel,Anne-Valerie Gendrel,Anwyn Apedaile,Heather Coker,Ausma Termanis,Ilona Zvetkova,Jonathan Godwin,Y. Amy Tang,Derek Huntley,Giovanni Montana,Steven Taylor,Eleni Giannoulatou,Edith Heard,Irina Stancheva,Neil Brockdorff +14 more
TL;DR: It is shown that inactive X chromosome (Xi) CGI methylation can occur via either Smchd1-dependent or -independent pathways, which differ with respect to their sequence characteristics and immediate chromosomal environment.
References
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Book
Antibodies: A Laboratory Manual
Ed Harlow,David P. Lane +1 more
TL;DR: A second edition of Antibodies: A Laboratory Manual is being published in September 2013, Revised, extended and updated by Edward Greenfield of the Dana-Farber Cancer Center, the material has been recast with extensive new information and new chapters have been added.
Book
Manipulating the mouse embryo: A laboratory manual
TL;DR: Here are recorded the tech- niques for preparing, inserting and analysing DNA sequences, for retroviral infection of mice, for production and use of EC and EK cells as vehicles for engineered sequences and for nuclear transplantation - all against a background of the basic procedures required for pro- ducing and handling the em- bryos.
Journal ArticleDOI
Gene Action in the X -chromosome of the Mouse ( Mus musculus L.)
TL;DR: Ohno and Hauschka1 showed that in female mice one chromosome of mammary carcinoma cells and of normal diploid cells of the ovary, mammary gland and liver was heteropyKnotic and suggested that the so-called sex chromatin was composed of one heteropyknotic X-chromosome.
Journal ArticleDOI
Histone methyltransferase activity associated with a human multiprotein complex containing the Enhancer of Zeste protein
TL;DR: The isolation of a multiprotein E(z) complex that contains extra sex combs, suppressor of zeste-12, and the histone binding proteins RbAp46/RbAp48 is reported, which possesses HMT activity with specificity for Lys 9 (K9) and Lys 27 (K27) of histone H3.
Book
Teratocarcinomas and embryonic stem cells : a practical approach
TL;DR: The results allowed us to assess the importance of knowing the carrier and removal status of canine coronavirus, as a source of infection for other animals, not necessarily belonging to the same breeds.