PRDMI14による始原生殖細胞特異的なエピゲノム調節とその機能

関西学院大

Tuning of Sry expression by H3K9 methylation and demethylation

Histone H3 lysine 9 (H3K9) methylation is a hallmark of heterochromatin. H3K9 demethylation is crucial in mouse sex determination; The H3K9 demethylase Jmjd1a deficiency leads to increased H3K9 methylation at the Sry locus in embryonic gonads, thereby compromising Sry expression and causing male-to-female sex reversal. We hypothesized that the H3K9 methylation level at the Sry locus is finely tuned by the balance in activities between the H3K9 demethylase Jmjd1a and an unidentified H3K9 methyltransferase to ensure correct Sry expression. Here we identified the GLP/G9a H3K9 methyltransferase complex as the enzyme catalyzing H3K9 methylation at the Sry locus. Based on this finding, we tried to rescue the sex-reversal phenotype of Jmjd1a-deficient mice by modulating GLP/G9a complex activity. A heterozygous GLP mutation rescued the sex-reversal phenotype of Jmjd1a-deficient mice by restoring Sry expression. The administration of a chemical inhibitor of GLP/G9a enzyme into Jmjd1a-deficient embryos also successfully rescued sex reversal. Our study not only reveals the molecular mechanism underlying the tuning of Sry expression but also provides proof on the principle of therapeutic strategies based on the pharmacological modulation of epigenetic balance

PRDM14 Drives OCT3/4 Recruitment via Active Demethylation in the Transition from Primed to Naive Pluripotency

Primordial germ cells (PGCs) are specified from epiblast cells in mice. Genes associated with naive pluripotency are repressed in the transition from inner cell mass to epiblast cells, followed by upregulation after PGC specification. However, the molecular mechanisms underlying the reactivation of pluripotency genes are poorly characterized. Here, we exploited the in vitro differentiation of epiblast-like cells (EpiLCs) from embryonic stem cells (ESCs) to elucidate the molecular and epigenetic functions of PR domain-containing 14 (PRDM14). We found that Prdm14 overexpression in EpiLCs induced their conversion to ESC-like cells even in the absence of leukemia inhibitory factor in adherent culture. This was impaired by the loss of Kruppel-like factor 2 and ten-eleven translocation (TET) proteins. Furthermore, PRDM14 recruited OCT3/4 to the enhancer regions of naive pluripotency genes via TET-base excision repair-mediated demethylation. Our results provide evidence that PRDM14 establishes a transcriptional network for naive pluripotency via active DNA demethylation

Combined Loss of JMJD1A and JMJD1B Reveals Critical Roles for H3K9 Demethylation in the Maintenance of Embryonic Stem Cells and Early Embryogenesis

Histone H3 lysine 9 (H3K9) methylation is unevenly distributed in mammalian chromosomes. However, the molecular mechanism controlling the uneven distribution and its biological significance remain to be elucidated. Here, we show that JMJD1A and JMJD1B preferentially target H3K9 demethylation of gene-dense regions of chromosomes, thereby establishing an H3K9 hypomethylation state in euchromatin. JMJD1A/JMJD1B-deficient embryos died soon after implantation accompanying epiblast cell death. Furthermore, combined loss of JMJD1A and JMJD1B caused perturbed expression of metabolic genes and rapid cell death in embryonic stem cells (ESCs). These results indicate that JMJD1A/JMJD1B-meditated H3K9 demethylation has critical roles for early embryogenesis and ESC maintenance. Finally, genetic rescue experiments clarified that H3K9 overmethylation by G9A was the cause of the cell death and perturbed gene expression of JMJD1A/JMJD1B-depleted ESCs. We summarized that JMJD1A and JMJD1B, in combination, ensure early embryogenesis and ESC viability by establishing the correct H3K9 methylated epigenome

The mouse Sry locus harbors a cryptic exon that is essential for male sex determination

The mammalian sex-determining gene induces male development. Since its discovery 30 years ago, has been believed to be a single-exon gene. Here, we identified a cryptic second exon of mouse and a corresponding two-exon type () transcript. XY mice lacking were sex-reversed, and ectopic expression of in XX mice induced male development. messenger RNA is expressed similarly to that of canonical single-exon type (), but SRY-T protein is expressed predominantly because of the absence of a degron in the C terminus of SRY-S. exon2 appears to have evolved recently in mice through acquisition of a retrotransposon-derived coding sequence to replace the degron. Our findings suggest that in nature, SRY-T, not SRY-S, is the bona fide testis-determining factor

Combined Loss of JMJD1A and JMJD1B Reveals Critical Roles for H3K9 Demethylation in the Maintenance of Embryonic Stem Cells and Early Embryogenesis

Histone H3 lysine 9 (H3K9) methylation is unevenly distributed in mammalian chromosomes. However, the molecular mechanism controlling the uneven distribution and its biological significance remain to be elucidated. Here, we show that JMJD1A and JMJD1B preferentially target H3K9 demethylation of gene-dense regions of chromosomes, thereby establishing an H3K9 hypomethylation state in euchromatin. JMJD1A/JMJD1B-deficient embryos died soon after implantation accompanying epiblast cell death. Furthermore, combined loss of JMJD1A and JMJD1B caused perturbed expression of metabolic genes and rapid cell death in embryonic stem cells (ESCs). These results indicate that JMJD1A/JMJD1B-meditated H3K9 demethylation has critical roles for early embryogenesis and ESC maintenance. Finally, genetic rescue experiments clarified that H3K9 overmethylation by G9A was the cause of the cell death and perturbed gene expression of JMJD1A/JMJD1B-depleted ESCs. We summarized that JMJD1A and JMJD1B, in combination, ensure early embryogenesis and ESC viability by establishing the correct H3K9 methylated epigenome

Rescuing the aberrant sex development of H3K9 demethylase Jmjd1a-deficient mice by modulating H3K9 methylation balance

<div><p>Histone H3 lysine 9 (H3K9) methylation is a hallmark of heterochromatin. H3K9 demethylation is crucial in mouse sex determination; The H3K9 demethylase Jmjd1a deficiency leads to increased H3K9 methylation at the <i>Sry</i> locus in embryonic gonads, thereby compromising <i>Sry</i> expression and causing male-to-female sex reversal. We hypothesized that the H3K9 methylation level at the <i>Sry</i> locus is finely tuned by the balance in activities between the H3K9 demethylase Jmjd1a and an unidentified H3K9 methyltransferase to ensure correct <i>Sry</i> expression. Here we identified the GLP/G9a H3K9 methyltransferase complex as the enzyme catalyzing H3K9 methylation at the <i>Sry</i> locus. Based on this finding, we tried to rescue the sex-reversal phenotype of Jmjd1a-deficient mice by modulating GLP/G9a complex activity. A heterozygous <i>GLP</i> mutation rescued the sex-reversal phenotype of Jmjd1a-deficient mice by restoring <i>Sry</i> expression. The administration of a chemical inhibitor of GLP/G9a enzyme into Jmjd1a-deficient embryos also successfully rescued sex reversal. Our study not only reveals the molecular mechanism underlying the tuning of <i>Sry</i> expression but also provides proof on the principle of therapeutic strategies based on the pharmacological modulation of epigenetic balance.</p></div

The <i>GLP</i> mutation rescues the reduced expression of Sry in Jmjd1a-deficient embryos.

<p>(A, C) Co-immunostaining profiles of Sry (A) and Sox9 (C) with a gonadal somatic cell marker, Gata4, in the center regions of E11.5 gonads of the indicated genotypes. Scale bar, 20 μm. (B, D) The ratios of cells positive for Sry (B) and Sox9 (D) to Gata4-positive cells in E11.5 gonads of the indicated genotypes. Numbers of examined animals are shown above the bars. All data are presented as mean ± SD. * <i>P</i> < 0.05; ** <i>P</i> < 0.01.</p

The <i>GLP</i> mutation rescues abnormal sex differentiation of XY Jmjd1a-deficient embryos.

<p>(A) Evaluation of sex differentiation of E13.5 embryonic gonads by immunofluorescence analysis. Sox9 and Foxl2 mark testicular Sertoli and ovarian somatic cells, respectively. The enlarged box indicates that the <i>GLP</i> mutation rescued the tubule-like structure that was absent in XY <i>Jmjd1a</i>^Δ/Δ gonads. Scale bar, 200 μm. (B) Quantification of Sox9- and Foxl2-positive cells in E13.5 gonads of the indicated genitypes. Numbers of embryos examined are shown above the bars. Data are presented as mean ± SD. * <i>P</i> < 0.05; ** <i>P</i> < 0.01.</p