Methylation of cytosines (5meC) is a widespread heritable DNA modification. During mammalian development, two global demethylation events are followed by waves of de novo DNA methylation. In vivo mechanisms of DNA methylation establishment are largely uncharacterized. Here, we use Saccharomyces cerevisiae as a system lacking DNA methylation to define the chromatin features influencing the activity of the murine DNMT3B. Our data demonstrate that DNMT3B and H3K4 methylation are mutually exclusive and that DNMT3B is co-localized with H3K36 methylated regions. In support of this observation, DNA methylation analysis in yeast strains without Set1 and Set2 shows an increase of relative 5meC levels at the transcription start site and a decrease in the gene-body, respectively. We extend our observation to the murine male germline, where H3K4me3 is strongly anti-correlated while H3K36me3 correlates with accelerated DNA methylation. These results show the importance of H3K36 methylation for gene-body DNA methylation in vivo.DOI: http://dx.doi.org/10.7554/eLife.06205.001

The heart switches its energy substrate from glucose to fatty acids at birth, and maternal hyperglycemia is associated with congenital heart disease. However, little is known about how blood glucose impacts heart formation. Using a chemically defined human pluripotent stem-cell-derived cardiomyocyte differentiation system, we found that high glucose inhibits the maturation of cardiomyocytes at genetic, structural, metabolic, electrophysiological, and biomechanical levels by promoting nucleotide biosynthesis through the pentose phosphate pathway. Blood glucose level in embryos is stable in utero during normal pregnancy, but glucose uptake by fetal cardiac tissue is drastically reduced in late gestational stages. In a murine model of diabetic pregnancy, fetal hearts showed cardiomyopathy with increased mitotic activity and decreased maturity. These data suggest that high glucose suppresses cardiac maturation, providing a possible mechanistic basis for congenital heart disease in diabetic pregnancy.

The organization of nucleosomes influences transcriptional activity by controlling accessibility of DNA binding proteins to the genome. Genome-wide nucleosome binding profiles have identified a canonical nucleosome organization at gene promoters, where arrays of well-positioned nucleosomes emanate from nucleosome-depleted regions. The mechanisms of formation and the function of canonical promoter nucleosome organization remain unclear. Here we analyze the genome-wide location of nucleosomes during zebrafish embryogenesis and show that well-positioned nucleosome arrays appear on thousands of promoters during the activation of the zygotic genome. The formation of canonical promoter nucleosome organization is independent of DNA sequence preference, transcriptional elongation, and robust RNA polymerase II (Pol II) binding. Instead, canonical promoter nucleosome organization correlates with the presence of histone H3 lysine 4 trimethylation (H3K4me3) and affects future transcriptional activation. These findings reveal that genome activation is central to the organization of nucleosome arrays during early embryogenesis.

Transgenic mice that overexpress metallothionein-I (MT-I) accumulate mo re MT-I and zinc in major organs than do control mice. The effects of overexpression of MT-I on resistance to dietary zinc deficiency were examined by feeding transgenic and control mice a zinc-deficient (0.5-1.5 micron/gram) or a zinc adequate (50 micron/g) diet and by measuring effects on pregnancy. When pregnant mice were maintained under conditions of dietary zinc deficiency, the number of resorptions and teratogenic defects of fetuses was greatly reduced in transgenic compared with control mice. Differences between transgenic and controls were not apparent at d 8 of pregnancy (d 1 = vaginal plug) but were apparent by d 14. This result suggests that the larger maternal zinc pool in the transgenic females allows fetal development to progress normally for a longer period of time. However, neither transgenic nor control zinc-deficient mice could complete pregnancy. Pancreatic MT concentrations were the greatest in zinc adequate transgenic mice. Moreover, there was >10-fold more MT per gram wet weight in the pancreas of transgenic mice than in any other organ examined. Pancreatic MT concentrations were an exceptionally sensitive indicator of zinc deficiency. Pancreatic MT declined 99.8% and zinc declined to basal levels by d 14 of pregnancy when transgenic and control mice were fed a zinc-deficient diet, whereas MT concentrations in other organs decreased only modestly. We suggest that the larger pool of zinc MT in the transgenic mice provides a biologically important labile pool of zinc during periods of zinc deficiency.

SummaryTo determine the role for mutations of MECP2 in Rett syndrome, we generated isogenic lines of human induced pluripotent stem cells, neural progenitor cells, and neurons from patient fibroblasts with and without MECP2 expression in an attempt to recapitulate disease phenotypes in vitro. Molecular profiling uncovered neuronal-specific gene expression changes, including induction of a senescence-associated secretory phenotype (SASP) program. Patient-derived neurons made without MECP2 showed signs of stress, including induction of P53, and senescence. The induction of P53 appeared to affect dendritic branching in Rett neurons, as P53 inhibition restored dendritic complexity. The induction of P53 targets was also detectable in analyses of human Rett patient brain, suggesting that this disease-in-a-dish model can provide relevant insights into the human disorder.

Oxidative stress and the inflammatory response may play roles in the pathogenesis of acute pancreatitis. Herein, we characterized pancreatic expression of oxidative stress-responsive genes [c-fos, heme oxygenase-1 (HO-1), and metallothionein-I (MT-I)] and cytokine genes [interleukin-1 beta (IL-1 beta), IL-6, and tumor necrosis factor-alpha (TNF-alpha)] during caerulein-induced acute pancreatitis in the mouse. c-fos, HO-1, and MT-I mRNAs were coordinately and rapidly (3-7 h) upregulated, and HO-1 and MT-I protein levels were increased slightly in the pancreas during acute pancreatitis. In addition, IL-1 beta, IL-6, and TNF-alpha mRNAs were rapidly (7 h) upregulated in the pancreas, and intrapancreatic IL-1 beta and IL-6 protein levels rapidly increased (3-fold and 6.4-fold, respectively) during acute pancreatitis. These studies suggest that oxidative stress and inflammation each occur in the pancreas during the early stages of acute pancreatitis. However, under a limited set of experimental conditions, we found that an insult that causes pancreatic oxidative stress (diethylmaleate) or one that induces an inflammatory response (bacterial lipopolysaccharide), or a combination of these agents, did not cause the changes characteristic of acute pancreatitis. Therefore, simply inducing oxidative stress and/or inflammation may be insufficient to initiate acute pancreatitis.

Taken together, these findings demonstrate that simple topological changes in cardiac fibroblast organization are sufficient to induce chromatin remodeling and global changes in gene expression with potential functional consequences for the healing heart.

The mouse metallothionein (MT) gene family consists of four known members (MT‐I through IV) clustered on chromosome 8. Studies reported herein examine the expression and regulation of the MT‐III and MT‐IV genes in specific cell types in the maternal reproductive tract, developing embryo, and fetus known to express the MT‐I and ‐II genes. MT‐III and MT‐IV mRNAs were absent from the visceral yolk sac, placenta, and fetal liver, tissues with high levels of MT‐I and MT‐II mRNAs. In contrast, MT‐III and MT‐IV mRNAs were both abundant in the maternal deciduum, and in experimentally induced deciduoma on 7 and 8 days postcoitum (1 dpc = vaginal plug), as are MT‐I and ‐II mRNAs. The abundance of each of these MT mRNAs increased coordinately during development of the deciduum (6–8 dpc), and in situ hybridization localized MT‐I, MT‐III, and MT‐IV mRNAs to the secondary decidual zone of the antimesometrial region on 8 dpc, where in some regions all of the cells were apparently positive. Thus, all of the known mouse MT genes are co‐expressed in at least some of the cells in the secondary decidual zone. Electrophoretic analysis of decidual MT suggested that the MT‐I, ‐II, and ‐III isoforms are abundant proteins in the secondary deciduum. Bacterial endotoxin‐lipopolysaccharide (LPS) and Zn are powerful inducers of MT‐I and MT‐II gene expression in many adult organs, whereas these agents apparently have little effect on MT‐III and MT‐IV gene expression. Neither of these agents significantly effected levels of decidual MT‐III or MT‐IV mRNAs in vivo or in primary cultures of decidual cells in vitro, and only modest effects of Zn on MT‐I mRNA levels were noted. During 2 days of in vitro culture, decidual cell MT‐I and MT‐III mRNA levels remained elevated while MT‐IV mRNA levels decreased. Thus, expression of the mouse MT gene locus in the deciduum appears to be developmentally regulated, and in this tissue, the MT genes are refractory to induction by Zn or inflammation. © 1996 Wiley‐Liss, Inc.