Chromatin dynamics and the role of G9a in gene regulation and enhancer silencing during early mouse development.

Early mouse development is accompanied by dynamic changes in chromatin modifications, including G9a-mediated histone H3 lysine 9 dimethylation (H3K9me2), which is essential for embryonic development. Here we show that genome-wide accumulation of H3K9me2 is crucial for postimplantation development, and coincides with redistribution of enhancer of zeste homolog 2 (EZH2)-dependent histone H3 lysine 27 trimethylation (H3K27me3). Loss of G9a or EZH2 results in upregulation of distinct gene sets involved in cell cycle regulation, germline development and embryogenesis. Notably, the H3K9me2 modification extends to active enhancer elements where it promotes developmentally-linked gene silencing and directly marks promoters and gene bodies. This epigenetic mechanism is important for priming gene regulatory networks for critical cell fate decisions in rapidly proliferating postimplantation epiblast cells.Wellcome Trust: Jan J Zylicz, Ufuk Günesdogan, Jamie A Hackett, Delphine Cougot, Caroline Lee, MA Surani, WT096738; European Commission (EC): Ufuk Günesdogan; Wellcome Trust: Jan J Zylicz, RG44593This is the final version of the article. It was first available from eLife via http://dx.doi.org/10.7554/eLife.0957

G9a regulates temporal preimplantation developmental program and lineage segregation in blastocyst

Early mouse development is regulated and accompanied by dynamic changes in chromatin modifications, including G9a-mediated histone H3 lysine 9 dimethylation (H3K9me2). Previously, we provided insights into its role in post-implantation development (Zylicz et al., 2015). Here we explore the impact of depleting the maternally inherited G9a in oocytes on development shortly after fertilisation. We show that G9a accumulates typically at 4 to 8-cell stage to promote timely repression of a subset of 4-cell stage-specific genes. Loss of maternal inheritance of G9a disrupts the gene regulatory network resulting in developmental delay and destabilisation of inner cell mass lineages by the late blastocyst stage. Our results indicate a vital role of this maternally inherited epigenetic regulator in creating conducive conditions for developmental progression and on cell fate choices.Wellcome Trust/CRUK Gurdon Institute, University of Cambridge Wellcome Trust/Medical Research Council, Stem Cell Institute, University of Cambridg

PRMT5 protects genomic integrity during global DNA demethylation in primordial germ cells and preimplantation embryos.

Primordial germ cells (PGCs) and preimplantation embryos undergo epigenetic reprogramming, which includes comprehensive DNA demethylation. We found that PRMT5, an arginine methyltransferase, translocates from the cytoplasm to the nucleus during this process. Here we show that conditional loss of PRMT5 in early PGCs causes complete male and female sterility, preceded by the upregulation of LINE1 and IAP transposons as well as activation of a DNA damage response. Similarly, loss of maternal-zygotic PRMT5 also leads to IAP upregulation. PRMT5 is necessary for the repressive H2A/H4R3me2s chromatin modification on LINE1 and IAP transposons in PGCs, directly implicating this modification in transposon silencing during DNA hypomethylation. PRMT5 translocates back to the cytoplasm subsequently, to participate in the previously described PIWI-interacting RNA (piRNA) pathway that promotes transposon silencing via de novo DNA remethylation. Thus, PRMT5 is directly involved in genome defense during preimplantation development and in PGCs at the time of global DNA demethylation.U.G. was supported by a Marie Sk1odowska Curie Intra-European Fellowship. J.J.Z. was a recipient of a Wellcome Trust PhD Studentship (RG44593). This research was supported by grants from the Wellcome Trust to M.A.S. (WT096738).This is the final published version. It first appeared at http://www.cell.com/molecular-cell/abstract/S1097-2765%2814%2900787-4

NANOG alone induces germ cells in primed epiblast in vitro by activation of enhancers.

Nanog, a core pluripotency factor in the inner cell mass of blastocysts, is also expressed in unipotent primordial germ cells (PGCs) in mice, where its precise role is yet unclear. We investigated this in an in vitro model, in which naive pluripotent embryonic stem (ES) cells cultured in basic fibroblast growth factor (bFGF) and activin A develop as epiblast-like cells (EpiLCs) and gain competence for a PGC-like fate. Consequently, bone morphogenetic protein 4 (BMP4), or ectopic expression of key germline transcription factors Prdm1, Prdm14 and Tfap2c, directly induce PGC-like cells (PGCLCs) in EpiLCs, but not in ES cells. Here we report an unexpected discovery that Nanog alone can induce PGCLCs in EpiLCs, independently of BMP4. We propose that after the dissolution of the naive ES-cell pluripotency network during establishment of EpiLCs, the epigenome is reset for cell fate determination. Indeed, we found genome-wide changes in NANOG-binding patterns between ES cells and EpiLCs, indicating epigenetic resetting of regulatory elements. Accordingly, we show that NANOG can bind and activate enhancers of Prdm1 and Prdm14 in EpiLCs in vitro; BLIMP1 (encoded by Prdm1) then directly induces Tfap2c. Furthermore, while SOX2 and NANOG promote the pluripotent state in ES cells, they show contrasting roles in EpiLCs, as Sox2 specifically represses PGCLC induction by Nanog. This study demonstrates a broadly applicable mechanistic principle for how cells acquire competence for cell fate determination, resulting in the context-dependent roles of key transcription factors during development.This is the author accepted manuscript. The final version is available from Nature Publishing Group via http://dx.doi.org/10.1038/nature1648

SPEN integrates transcriptional and epigenetic control of X-inactivation

Xist represents a paradigm for the function of long non-coding RNA in epigenetic regulation, although how it mediates X-chromosome inactivation (XCI) remains largely unexplained. Several proteins that bind to Xist RNA have recently been identified, including the transcriptional repressor SPEN(1-3), the loss of which has been associated with deficient XCI at multiple loci(2-6). Here we show in mice that SPEN is a key orchestrator of XCI in vivo and we elucidate its mechanism of action. We show that SPEN is essential for initiating gene silencing on the X chromosome in preimplantation mouse embryos and in embryonic stem cells. SPEN is dispensable for maintenance of XCI in neural progenitors, although it significantly decreases the expression of genes that escape XCI. We show that SPEN is immediately recruited to the X chromosome upon the upregulation of Xist, and is targeted to enhancers and promoters of active genes. SPEN rapidly disengages from chromatin upon gene silencing, suggesting that active transcription is required to tether SPEN to chromatin. We define the SPOC domain as a major effector of the gene-silencing function of SPEN, and show that tethering SPOC to Xist RNA is sufficient to mediate gene silencing. We identify the protein partners of SPOC, including NCoR/SMRT, the m(6)A RNA methylation machinery, the NuRD complex, RNA polymerase II and factors involved in the regulation of transcription initiation and elongation. We propose that SPEN acts as a molecular integrator for the initiation of XCI, bridging Xist RNA with the transcription machinery-as well as with nucleosome remodellers and histone deacetylases-at active enhancers and promoters

Testosterone increases amygdala reactivity in middle-aged women to a young adulthood level

Contains fulltext : 77294.pdf (publisher's version ) (Closed access)Testosterone modulates mood and sexual function in women. However, androgen levels decline with age, which may relate to the age-associated change in sexual functioning and the prevalence of mood and anxiety disorders. These effects of testosterone are potentially mediated by the amygdala. In the present study, we investigated whether the age-related decline in androgen levels is associated with reduced amygdala activity, and whether exogenous testosterone can restore amygdala activity. Healthy young and middle-aged women participated during the early follicular phase of the menstrual cycle, and amygdala responses to biologically salient stimuli were measured with functional magnetic resonance imaging (fMRI). Androgen levels were lower in middle-aged than young women, which was associated with decreased amygdala reactivity. Endogenous testosterone levels correlated positively with amygdala reactivity across the young and middle-aged women. The middle-aged women received a single nasal dose of testosterone in a double-blind, placebo-controlled, crossover manner, which rapidly increased amygdala reactivity to a level comparable to the young women. The enhanced testosterone levels correlated positively with superior frontal cortex responses and negatively with orbitofrontal cortex responses across individuals, which may reflect testosterone-induced changes in amygdala regulation. These results show that testosterone modulates amygdala reactivity in women, and suggest that the age-related decline in androgen levels contribute to the decrease in amygdala reactivity