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  • Molecular Cell


    tic W


    e hrd ou in

    et al. (2012) chart the dynamics of DNA

    rior epiblast by external signals around

    A second key observation is that while

    further exploration.

    sine to 5-hydroxymethylcytosine. Tet3

    high-throughput sequencing, a methodment in the gonads occurs from e12.5.

    Seisenberger et al. (2012) optimized

    bisulfite conversion coupled to next-

    methylation dynamics from fertiliza-

    tion to the e7.5 epiblast (Figure 1).

    Using Reduced Representation BS-Seq

    assessment of methylation. Many CpG

    dinucleotides hemimethylated in e9.5

    PGCs were completely unmethylatedgeneration sequencing (BS-Seq) for small

    input quantities, enabling assessment at

    (RRBS), which informs on DNA methyla-

    tion at base pair resolution but does not

    by e13.5. Moreover, in hemimethylated

    sequences, the methylated CpGs weree6.5. From e9.5, the cells migrate to the

    genital ridge, and sex-specific develop-

    This study complements recent work

    by Smith et al. (2012) which explores

    that keeps double-stranded DNA to-

    gether and permits a strand-specificmethylation during PGC development,

    revealing insights into reprogramming

    mechanisms and demonstrating that

    some sequences can be resistant to

    demethylation, providing the strongest

    candidates yet for carriers of trans-

    generational epigenetic inheritance in


    Murine PGCs are induced in the poste-

    reprogramming is linked to transient

    activation of the pluripotency gene net-

    work, global demethylation is not coupled

    to promiscuous transcriptional activity.

    This implies that other mechanisms,

    likely histone modifications, play the lead

    role in PGC transcriptional control. The

    nature of these mechanisms, and their

    relevance in other contexts, warrants

    protein is not detected in PGCs, sug-

    gesting the involvement of alternative

    mechanisms (Hackett et al., 2012). Sei-

    senberger et al. (2012) provide strong

    evidence that PGC demethylation is pre-

    dominantly passive, meaning that methy-

    lation is not maintained during DNA

    replication. To demonstrate this, the

    authors make use of hairpin bisulfitePreviews

    Resetting for the N

    Michael Cowley1 and Rebecca J. Oake1Department of Medical and Molecular Gene*Correspondence:

    In this issue of Molecular Cell, Seisepigenetic reprogramming of primootency, and identifying potential ro

    The totipotent zygote gives rise to all of

    the differentiated cell types of the adult

    by embarking on a program of cell division

    and specification that progressively limits

    cellular potential. Epigenetic marks, in-

    cluding DNA methylation, are critical for

    mediating this program. At fertilization,

    the genome undergoes global methy-

    lation erasure, creating a blank canvas

    onto which epigenetic profiles can be

    painted as somatic cell lineages are

    specified. However, one fundamental

    question is this: what happens to the

    cells that will form the germline of the

    developing embryo? Primordial germ

    cells (PGCs) must achieve an epigenetic

    state distinct from somatic cells, retaining

    the plasticity necessary for producing

    gametes and ultimately a totipotent

    zygote in the next generation (Hackett

    et al., 2012). Since PGCs are derived

    from cells already primed for a somatic

    fate, they must be reprogrammed by

    another wave of demethylation. This is

    crucial for ensuring that epigenetic infor-

    mation is reset in each generation. In this

    issue of Molecular Cell, Seisenbergerext Generation

    ,*s, Kings College London, Eighth Floor Tower

    nberger et al. (2012) refine DNA metial germ cells, defining the timingstes to transgenerational epigenetic

    base pair resolution of DNA methylation

    in e6.5 epiblasts (selected to represent

    the ground state prior to PGC specifi-

    cation) plus PGCs isolated from later

    developmental stages. The first striking

    finding is the dramatically reduced global

    levels of DNA methylation in e9.5 PGCs

    comparedwith thee6.5 epiblast, revealing

    that most erasure occurs within this

    developmental window (Figure 1). Some

    specific sequences reproducibly escape

    this initial loss of methylation but become

    demethylated after e10.5. These late

    demethylaters include the CpG islands

    associated with differentially methylated

    regions (DMRs) of imprinted loci. It is

    noteworthy that DMRs escape methy-

    lation erasure altogether during the post-

    fertilization demethylation wave (Figure 1),

    and the authors suggest that their

    delayed demethylation in PGCs might

    arise from the persistence of this protec-

    tive mechanism, involving binding of the

    zinc finger protein Zfp57 (Li et al., 2008).

    However, the dynamics of Zfp57 activity,

    and how these might be coupled to re-

    programming in PGCs, remain unclear.Molecular Cell 48, Ding, Guys Hospital, London SE1 9RT, UK

    ylation mapping to interrogate thef methylation loss, linking to plurip-heritance.

    provide complete genome coverage,

    Smith et al. (2012) confirmed two key

    transitions in early mouse development:

    first, the rapid methylation erasure of

    the paternal genome and the passive

    erasure of the maternal genome after

    fertilization; and second, the genome-

    wide gain of methylation between the

    inner cell mass of the blastocyst and the

    early epiblast. These are the marks that

    must then be reset in PGCs. Smallwood

    et al. (2011) also exploited RRBS to

    interrogate methylation dynamics during

    oocyte development. Together, these

    studies enable us to build a compre-

    hensive picture of the timings of methy-

    lation dynamics during development

    that underpin totipotency and differentia-

    tion (Figure 1).

    What is the mechanism for epigenetic

    reprogramming in PGCs? This is a

    particularly hot topic because of the

    recent discovery that Tet3 orchestrates

    genome-wide demethylation of the

    paternal genome upon fertilization (Gu

    et al., 2011). This is an active mechanism

    involving the oxidation of 5-methylcyto-ecember 28, 2012 2012 Elsevier Inc. 819

  • Molecular Cellpredominantly located on the same

    DNA strand, consistent with a failure to

    maintain methylation during DNA replica-

    tion. While this suggests demethylation

    in PGCs is passive, other studies have

    shown an involvement of active mecha-

    nisms (reviewed in Hackett et al., 2012).

    The relative importance of these pro-

    cesses requires clarification, but the po-

    tential redundancy underlines the funda-

    mental importance of reprogramming

    in ensuring the integrity of information


    Through the analysis of DNA methyla-

    tion during PGC development, Seisen-

    berger et al. (2012) assess resistance to

    genome-wide demethylation, identifying

    regions of both consistent and variable

    methylation. From this, the authors

    propose a potential mechanism for the

    much debated transgenerational epige-

    netic inheritance. This form of inheritance

    refers to non-sequence-based effects on

    phenotype passed from one generation

    Figure 1. DNA Methylation Dynamics during DAfter fertilization, the paternal genome (blue line) is deDifferentially methylated regions (DMRs) associatedpostimplantation (black line), but PGCs are not spece6.5, the figure shows the methylation dynamics inof sequences are late demethylaters and are not repare resistant to demethylation during both the postfPGC reprogramming, but their methylation status dugerm cells occurs, but the dynamics are sex specific. Mduring the growth phase, after birth. In adulthood, theShown below are the developmental windows examoocytes. GV, germinal vesicle oocytes. MII, metaphas

    820 Molecular Cell 48, December 28, 2012 to the next via the gametes (Daxinger

    and Whitelaw, 2012). The epigenetic

    carriers of this information must resist

    being cleared during early development

    and PGC reprogramming. It is known

    that intracisternal A particles (IAPs)

    remain methylated in mature gametes,

    preimplantation embryos, and PGCs

    (Popp et al., 2010), but this new study

    catalogs not only sequences that fully

    resist reprogramming but also some

    that variably escape. Intriguingly, the

    latter include some CpG islands (CGIs)

    located away from the influence of IAPs,

    termed variably erased CGIs (VECs), that

    exhibit variable erasure across multiple

    stages of development including mature

    gametes. These VECs present potential

    vehicles for the transmission of epige-

    netic information across generations,

    and could provide the mechanistic basis

    for the observed transgenerational inheri-

    tance of metabolic phenotypes, such as

    obesity, in animal models of nutritional

    evelopmental Reprogrammingmethylated rapidly by active mechanisms, while the mawith imprinted genes are protected from this erasureified until the epiblast stage (shading at top of figurethe cells that form the germline only. Most sequencrogrammed until after PGC migration. These includeertilization and the PGC reprogramming waves. Variaring postfertilization reprogramming is unclear. Followethylation is completed in prospermatogonia before b