Myst family genes encode lysine acetyltransferases that mainly mediate histone acetylation to control transcription, DNA replication and DNA damage response. They form tetrameric complexes with ...PHD-finger proteins (Brpfs or Jades) and small non-catalytic subunits Ing4/5 and Meaf6. Although all the components of the complex are well-conserved from yeast to mammals, the function of Meaf6 and its homologs has not been elucidated in any species. Here we revealed the role of Meaf6 utilizing inducible Meaf6 KO ES cells. By elimination of Meaf6, proliferation ceased although histone acetylations were largely unaffected. In the absence of Meaf6, one of the Myst family members Myst2/Kat7 increased the ability to interact with PHD-finger proteins. This study is the first indication of the function of Meaf6, which shows it is not essential for HAT activity but modulates the assembly of the Kat7 complex.
Genomic imprinting is regulated by differential methylation of the paternal and maternal genome. However, it remains unknown how parental imprinting is established during gametogenesis. In this ...study, we demonstrate that Dnmt3L, a protein sharing homology with DNA methyltransferases, Dnmt3a and Dnmt3b, but lacking enzymatic activity, is essential for the establishment of maternal methylation imprints and appropriate expression of maternally imprinted genes. We also show that Dnmt3L interacts with Dnmt3a and Dnmt3b and co-localizes with these enzymes in the nuclei of transfected cells, suggesting that Dnmt3L may regulate genomic imprinting via the Dnmt3 family enzymes. Consistent with this model, we show that Dnmt3a â/â , Dnmt3b +/â mice also fail to establish maternal methylation imprints. In addition, both Dnmt3a and Dnmt3L are required for spermatogenesis. Together, our findings suggest that Dnmt3L may cooperate with Dnmt3 family methyltransferases to carry out de novo methylation of maternally imprinted genes in oocytes.
Ten-eleven translocation (TET) proteins oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). 5fC and 5caC can be excised and ...repaired by the base excision repair (BER) pathway, implicating 5mC oxidation in active DNA demethylation. Genome-wide DNA methylation is erased in the transition from metastable states to the ground state of embryonic stem cells (ESCs) and in migrating primordial germ cells (PGCs), although some resistant regions become demethylated only in gonadal PGCs. Understanding the mechanisms underlying global hypomethylation in naive ESCs and developing PGCs will be useful for realizing cellular pluripotency and totipotency. In this study, we found that PRDM14, the PR domain-containing transcriptional regulator, accelerates the TET-BER cycle, resulting in the promotion of active DNA demethylation in ESCs. Induction of Prdm14 expression transiently elevated 5hmC, followed by the reduction of 5mC at pluripotency-associated genes, germline-specific genes and imprinted loci, but not across the entire genome, which resembles the second wave of DNA demethylation observed in gonadal PGCs. PRDM14 physically interacts with TET1 and TET2 and enhances the recruitment of TET1 and TET2 at target loci. Knockdown of TET1 and TET2 impaired transcriptional regulation and DNA demethylation by PRDM14. The repression of the BER pathway by administration of pharmacological inhibitors of APE1 and PARP1 and the knockdown of thymine DNA glycosylase (TDG) also impaired DNA demethylation by PRDM14. Furthermore, DNA demethylation induced by PRDM14 takes place normally in the presence of aphidicolin, which is an inhibitor of G1/S progression. Together, our analysis provides mechanistic insight into DNA demethylation in naive pluripotent stem cells and developing PGCs.
DNA methylation is an important epigenetic modification regulating various biological phenomena, including genomic imprinting and transposon silencing. It is known that methylation of the ...differentially methylated regions (DMRs) associated with paternally imprinted genes and of some repetitive elements occurs during male germ cell development in the mouse. We have performed a detailed methylation analysis of the paternally methylated DMRs (H19, Dlk1/Gtl2 and Rasgrf1), interspersed repeats SineB1, intracisternal A particle (IAP) and Line1 and satellite repeats (major and minor) to determine the timing of this de novo methylation in male germ cells. Furthermore, we have examined the roles of the de novo methyltransferases (Dnmt3a and Dnmt3b) and related protein (Dnmt3L) in this process. We found that methylation of all DMRs and repeats occurred progressively in fetal prospermatogonia and was completed by the newborn stage. Analysis of newborn prospermatogonia from germline-specific Dnmt3a and Dnmt3b knockout mice revealed that Dnmt3a mainly methylates the H19 and Dlk1/Gtl2 DMRs and a short interspersed repeat SineB1. Both Dnmt3a and Dnmt3b were involved in the methylation of Rasgrf1 DMR and long interspersed repeats IAP and Line1. Only Dnmt3b was required for the methylation of the satellite repeats. These results indicate both common and differential target specificities of Dnmt3a and Dnmt3b in vivo. Finally, all these sequences showed moderate to severe hypomethylation in Dnmt3L-deficient prospermatogonia, indicating the critical function and broad specificity of this factor in de novo methylation.
Low-modulus polypropylene (LMPP) with controlled stereoregularity was prepared based on the original catalyst technology Minami et al. (Polym J 47:227–34, 2015). The LMPP showed elastic recovery, ...while plastic deformation was dominant in isotactic polypropylene. To understand the underlying mechanism, changes in the morphology of LMPP under cyclic uniaxial elongation were investigated using in situ small-angle X-ray scattering (SAXS) and wide-angle X-ray diffraction (WAXD). During the first cycle, yielding appeared in the stress–strain (S–S) curve, and the residual strain was large. The SAXS pattern is a ring pattern at strain zero and then changes to a four-point pattern and a two-point pattern with increasing strain. This four-point pattern is derived from the undulating structure of the crystal lamellae. During the stretching process, this undulation is large, and fragmentation of the lamellae occurs. Therefore, the residual strain increases. During the second cycle, no yield appeared in the S–S curve, and the residual strain was small. The SAXS pattern changed quickly from a four-point pattern to a two-point pattern. This result suggests that little fragmentation of the lamellae occurs and that only the lamellae were rotating under elongation. Thus, plastic deformation hardly occurs, and the residual strain decreases. Based on these results, lamella fragmentation has a significant effect on the elastic-recovery rate.Low-modulus polypropylene (LMPP) with controlled stereoregularity showed elastic recovery. To understand the underlying mechanism, changes in the morphology of LMPP under cyclic uniaxial elongation were investigated using in situ SAXS and WAXD. During the first cycle, the undulating structure of the crystal lamellae is large, and fragmentation of the lamellae occurs. On the other hand, during the second cycle, the little fragmentation of the lamellae occurs and that only the lamellae were rotating under elongation. Based on these results, lamella fragmentation has a significant effect on the elastic-recovery rate.
Phase transitions of Mg2TiO4 and Fe2TiO4 were examined up to 28 GPa and 1600 °C using a multianvil apparatus. The quenched samples were examined by powder X-ray diffraction. With increasing pressure ...at high temperature, spinel-type Mg2TiO4 decomposes into MgO and ilmenite-type MgTiO3 which further transforms to perovskite-type MgTiO3. At ~21 GPa, the assemblage of MgTiO3 perovskite + MgO changes to 2MgO + TiO2 with baddeleyite (or orthorhombic I)-type structure. Fe2TiO4 undergoes transitions similar to Mg2TiO4 with pressure: spinel-type Fe2TiO4 dissociates into FeO and ilmenite-type FeTiO3 which transforms to perovskite-type FeTiO3. Both of MgTiO3 and FeTiO3 perovskites change to LiNbO3-type phases on release of pressure. In Fe2TiO4, however, perovskite-type FeTiO3 and FeO combine into calcium titanate-type Fe2TiO4 at ~15 GPa. The formation of calcium titanate-type Fe2TiO4 at high pressure may be explained by effects of crystal field stabilization and high spin–low spin transition in Fe2+ in the octahedral sites of calcium titanate-type Fe2TiO4. It is inferred from the determined phase relations that some of Fe2TiO4-rich titanomagnetite inclusions in diamonds recently found in São Luiz, Juina, Brazil, may be originally calcium titanate-type Fe2TiO4 at pressure above ~15 GPa in the transition zone or lower mantle and transformed to spinel-type in the upper mantle conditions.
The Polycomb group (PcG) proteins mediate heritable silencing of developmental regulators in metazoans, participating in one of two distinct multimeric protein complexes, the Polycomb repressive ...complexes 1 (PRC1) and 2 (PRC2). Although PRC2 has been shown to share target genes with the core transcription network, including Oct3/4, to maintain embryonic stem (ES) cells, it is still unclear whether PcG proteins and the core transcription network are functionally linked. Here, we identify an essential role for the core PRC1 components Ring1A/B in repressing developmental regulators in mouse ES cells and, thereby, in maintaining ES cell identity. A significant proportion of the PRC1 target genes are also repressed by Oct3/4. We demonstrate that engagement of PRC1 at target genes is Oct3/4-dependent, whereas engagement of Oct3/4 is PRC1-independent. Moreover, upon differentiation induced by Gata6 expression, most of the Ring1A/B target genes are derepressed and the binding of Ring1A/B to their target loci is also decreased. Collectively, these results indicate that Ring1A/B-mediated Polycomb silencing functions downstream of the core transcriptional regulatory circuitry to maintain ES cell identity.
De novo methylation of genomic DNA is a developmentally regulated process that is believed to play a pivotal role in regulation of genomic imprinting and X-chromosome inactivation in mammals. ...Aberrant de novo methylation of growth regulatory genes has been associated with tumorigenesis in humans. We have shown previously that de novo methylation persists in embryonic stem (ES) cells lacking Dnmt1, which encodes the constitutive DNA methyltransferase Dnmt1 (or MT1), indicating the existence of independently encoded de novo methyltransferases. We performed a TBLASTN search of the dbEST database using full-length bacterial type II cytosine-5 methyltransferase sequences as queries and found four matching human EST clones. Subsequent isolation and sequencing analysis of overlapping cDNA clones identified two homologous genes in both human and mouse which contain the highly conserved cytosine-5 methyltransferase motifs. The mouse genes are termed Dnmt3a and Dnmt3b as they show little sequence similarity to mouse Dnmt1 and Dnmt2 and masc1 from Ascobolus. Dnmt3a and Dnmt3b cDNA are 4,192 bp and 4,195 bp in length, encoding proteins of 908 and 859 amino acids, respectively. The Dnmt3b gene also encodes at least two shorter polypeptides of 840 and 777 amino-acid residues through alternative splicing. The human DNMT3A and DNMT3B cDNA are highly homologous to the mouse genes (data not shown).
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
DNA methylation regulates development and many epigenetic processes in mammals 1, and it is required for somatic cell growth and survival 2, 3. In contrast, embryonic stem (ES) cells can self-renew ...without DNA methylation 4–6. It remains unclear whether any lineage-committed cells can survive without DNA-methylation machineries. Unlike in somatic cells, DNA methylation is dispensable for imprinting and X-inactivation in the extraembryonic lineages 7–12. In ES cells, DNA methylation prevents differentiation into the trophectodermal fate 13. Here, we created triple-knockout (TKO) mouse embryos deficient for the active DNA methyltransferases Dnmt1, Dnmt3a, and Dnmt3b (TKO) by nuclear transfer (NT), and we examined their development. In chimeric TKO-NT and WT embryos, few TKO cells were found in the embryo proper, but they contributed to extraembryonic tissues. TKO ES cells showed increasing cell death during their differentiation into epiblast lineages, but not during differentiation into extraembryonic lineages. Furthermore, we successfully established trophoblastic stem cells (ntTS cells) from TKO-NT blastocysts. These TKO ntTS cells could self-renew, and they retained the fundamental gene expression patterns of stem cells. Our findings indicated that extraembryonic-lineage cells can survive and proliferate in the absence of DNA methyltransferases and that a cell's response to the stress of epigenomic damage is cell type dependent.
► NT embryos derived from Dnmt1/3a/3b-TKO ES cells can develop into blastocysts ► Dnmt1/3a/3b-TKO-NT derived cells can contribute to extraembryonic tissues in vivo ► Trophoblastic stem cells can self-renew in the absence of Dnmt1, Dnmt3a, and Dnmt3b ► Growth and survival defects resulting from the loss of Dnmts are lineage dependent
DNA methyltransferases Dnmt1, Dnmt3a and Dnmt3b cooperatively regulate cytosine methylation in CpG dinucleotides in mammalian genomes, providing an epigenetic basis for gene silencing and maintenance ...of genome integrity. Proper CpG methylation is required for the normal growth of various somatic cell types, indicating its essential role in the basic cellular function of mammalian cells. Previous studies using Dnmt1−/– or Dnmt3a−/–Dnmt3b−/– ES cells, however, have shown that undifferentiated embryonic stem (ES) cells can tolerate hypomethylation for their proliferation. In an attempt to investigate the effects of the complete loss of CpG DNA methyltransferase function, we established mouse ES cells lacking all three of these enzymes by gene targeting. Despite the absence of CpG methylation, as demonstrated by genome‐wide methylation analysis, these triple knockout (TKO) ES cells grew robustly and maintained their undifferentiated characteristics. TKO ES cells retained pericentromeric heterochromatin domains marked with methylation at Lys9 of histone H3 and heterochromatin protein‐1, and maintained their normal chromosome numbers. Our results indicate that ES cells can maintain stem cell properties and chromosomal stability in the absence of CpG methylation and CpG DNA methyltransferases.
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