DNA hypomethylation characterizes genes encoding tissue-dominant functional proteins in liver and skeletal muscle.

Each tissue has a dominant set of functional proteins required to mediate tissue-specific functions. Epigenetic modifications, transcription, and translational efficiency control tissue-dominant protein production. However, the coordination of these regulatory mechanisms to achieve such tissue-specific protein production remains unclear. Here, we analyzed the DNA methylome, transcriptome, and proteome in mouse liver and skeletal muscle. We found that DNA hypomethylation at promoter regions is globally associated with liver-dominant or skeletal muscle-dominant functional protein production within each tissue, as well as with genes encoding proteins involved in ubiquitous functions in both tissues. Thus, genes encoding liver-dominant proteins, such as those involved in glycolysis or gluconeogenesis, the urea cycle, complement and coagulation systems, enzymes of tryptophan metabolism, and cytochrome P450-related metabolism, were hypomethylated in the liver, whereas those encoding-skeletal muscle-dominant proteins, such as those involved in sarcomere organization, were hypomethylated in the skeletal muscle. Thus, DNA hypomethylation characterizes genes encoding tissue-dominant functional proteins.

Microarray transcriptome datasets of maternal-zygotic DNA methyltransferase 3aa -/- zebrafish during early developmental stages.

DNA methylation is an epigenetic regulator mediated by DNA methyltransferases (Dnmts). The methylation is involved in control of gene expression in vertebrates. It has been reported that there are mainly two types of de novo Dnmts, Dnmt3a and Dnmt3b, in mammals. These two Dnmts function in DNA methylation in the distinct or overlapping genomic regions. The zebrafish homologs of mammalian Dnmt3a are Dnmt3aa and Dnmt3ab. We generated a maternal-zygotic dnmt3aa deficient mutant (MZdnmt3aa) to identify the specific target regions for DNA methylation in the zebrafish genome and their function in the developmental process. Microarray analysis revealed alterations in gene expression by knock-out of dnmt3aa in early zebrafish development. Microarray datasets were produced from samples at five different developmental stages: 1-2 cell, shield, 5-somite, 1-day post fertilization (dpf), and 2 dpf. Herein, we present novel raw and processed transcriptome datasets generated by analysis of the MZdnmt3aa -/- mutant. The raw microarray data are available through the Gene Expression Omnibus (GEO), accession number GSE202646. These transcriptome data may be useful for comparing differences in gene expression among species of Dnmt3a mutants and for analyzing human diseases caused by DNMT3A such as acute myelogenous leukemia (AML).

Methylome data derived from maternal-zygotic DNA methyltransferase 3aa-/- zebrafish.

Genomic DNA methylation is an epigenetic marker mediated by DNA methyltransferases (Dnmts); in vertebrates, it comprises of a maintenance DNA methyltransferase, Dnmt1, and two de novo DNA methyltransferases (Dnmt3a and Dnmt3b). In zebrafish, there are two homologs of the mammalian Dnmt3a: Dnmt3aa and Dnmt3ab. A knockout (KO) mutant of zebrafish dnmt3aa was generated using the CRISPR/Cas9 genome-editing system as a new model for DNA methylation research. Since zebrafish dnmt3aa KO mutants were viable and fertile, a maternal-zygotic dnmt3aa deficient mutant (MZdnmt3aa) was generated. We performed whole-genome bisulfite sequencing (WGBS) to reveal the DNA methylation profile using this mutant and identified genomic regions with altered CpG methylation as differentially methylated regions (DMRs) in this mutant compared to those in the wild-type fish. We provided novel raw and processed datasets using the MZdnmt3aa KO mutant, and the raw data of WGBS are available through the Gene Expression Omnibus (GEO), accession number GSE178690.

Identification of aberrant transcription termination at specific gene loci with DNA hypomethylated transcription termination sites caused by DNA methyltransferase deficiency.

CpG methylation of genomic DNA is a well-known repressive epigenetic marker in eukaryotic transcription, and DNA methylation of promoter regions is correlated with gene silencing. In contrast to the promoter regions, the function of DNA methylation during transcription termination remains to be elucidated. A recent study revealed that mouse DNA methyltransferase 3a (Dnmt3a) mainly functions in de novo methylation in the promoter and gene body regions, including transcription termination sites (TTSs), during development. To investigate the relationship between DNA methylation overlapping the TTSs and transcription termination, we performed bioinformatics analysis using six pre-existing Dnmt-/- mouse cell datasets: four types of neurons (three Dnmt3a-/- and one Dnmt1-/- mutants) and two types of embryonic fibroblasts (MEFs) (Dnmt3a-/- and Dnmt3b-/- mutants). Combined analyses using methylome and transcriptome data revealed that read counts downstream of hypomethylated TTSs were increased in three types of neurons (two Dnmt3a-/- and one Dnmt1-/- mutants). Among these, an increase in chimeric transcripts downstream of the TTSs was observed in Dnmt3a-/- mature olfactory sensory neurons and Dnmt3a-/- agouti-related peptide (protein)-producing neurons, thereby indicating that read-through occurs in hypomethylated TTSs at specific gene loci in these two mutants. Conversely, in Dnmt3a-/- MEFs, we detected reductions in read counts downstream of hypomethylated TTSs. These results indicate that the hypomethylation of TTSs can both positively and negatively regulate transcription termination, dependent on Dnmt and cell types. This study is the first to identify the aberrant termination of transcription at specific gene loci with DNA hypomethylated TTSs attributable to Dnmt deficiency.

Compact and Scalable Large Vortex Array Generation Using Azocarbazole Polymer and Digital Hologram Printing Technique.

An integrated device capable of generating large number of multiplexed optical vortex beams with arbitrary topological charge is considered as one of the crucial requirement for driving information photonics forward. Here we report a simple method for simultaneous generation of 100 multiplexed optical vortex beams from a polymer film of size 1 mm2 and thickness of 30 µm. This is achieved through a combination of computer-generated holography, digital hologram printing and photoisomeric polymers. When the fabricated sample is illuminated with a collimated laser beam, a pre-determined vortex array with arbitrary topological charge is emitted. The polymer film easy to synthesize and exhibits a diffraction efficiency of 30% with a retention period longer than 50 days.

Field and laboratory metabolism and thermoregulation in rhinoceros auklets.

Seabirds spend most of their lives at sea, except when visiting their breeding sites. Since the thermal conductivity of water is 25 times higher than that of air, seabirds resting on water lose heat and expend a considerable amount of energy for thermoregulation. For example, rhinoceros auklet (Cerorhinca monocerata), a medium-sized (480620 g) alcid, spends most of its time floating on the sea. In order to estimate the cost of this behavior in terms of their daily energy expenditure (DEE), we studied rhinoceros auklets breeding on Teuri Island, Hokkaido Japan. We measured their resting metabolic rate (RMR) in air and on water by respirometry, and estimated their DEE by the doubly labeled water method. While RMR on water did not vary significantly between 10C and 15C, it was significantly higher at 5C. Air temperature (5.020.0C) had no effect on RMR. The DEE of free-ranging auklets averaged 1,005.5kJday1 (130.2, n=3). Our results indicate that RMRs are elevated for auklets resting on water, particularly below their lower critical temperature (LCT), compared with in air. Accordingly, spending time above their LCT on water at any time of year will provide enhanced benefits, particularly to seabirds such as rhinoceros auklets which rest a considerable amount of time on water.

The N-terminal domain of gastrulation brain homeobox 2 (Gbx2) is required for iridophore specification in zebrafish.

Although body color pattern formation by pigment cells plays critical roles in animals, pigment cell specification has not yet been fully elucidated. In zebrafish, there are three chromatophores: melanophore, iridophore, and xanthophore, that are derived from neural crest cells (NCCs). A recent study has reported the differentially expressed genes between melanophores and iridophores. Based on transcriptome data, we identified that Gbx2 is required for iridophore specification during development. In support of this, iridophore formation is suppressed by gbx2 knockdown by morpholino antisense oligonucleotide, at 72 h post fertilization (hpf) in zebrafish. Moreover, gbx2 is expressed in sox10-expressing NCCs and guanine crystal plates-containing iridophores during development at 24 and 48 hpf, respectively. In gbx2 knockdown zebrafish embryos, apoptosis of sox10-expressing NCCs was detected at 24 hpf without any effect on the formation of melanophores and xanthophores at 48 hpf. We further observed that the N-terminal domain of Gbx2 is able to rescue the iridophore formation defect caused by gbx2 knockdown. Our study provides insights into the requirement of N-terminal domain of Gbx2 for iridophore specification in zebrafish.

Seabird mortality induced by land-based artificial lights.

Artificial lights at night cause high mortality of seabirds, one of the most endangered groups of birds globally. Fledglings of burrow-nesting seabirds, and to a lesser extent adults, are attracted to and then grounded (i.e., forced to land) by lights when they fly at night. We reviewed the current state of knowledge of seabird attraction to light to identify information gaps and propose measures to address the problem. Although species in families such as Alcidae and Anatidae can be grounded by artificial light, the most affected seabirds are petrels and shearwaters (Procellariiformes). At least 56 species of Procellariiformes, more than one-third of them (24) threatened, are subject to grounding by lights. Seabirds grounded by lights have been found worldwide, mainly on oceanic islands but also at some continental locations. Petrel breeding grounds confined to formerly uninhabited islands are particularly at risk from light pollution due to tourism and urban sprawl. Where it is impractical to ban external lights, rescue programs of grounded birds offer the most immediate and employed mitigation to reduce the rate of light-induced mortality and save thousands of birds every year. These programs also provide useful information for seabird management. However, these data are typically fragmentary, biased, and uncertain and can lead to inaccurate impact estimates and poor understanding of the phenomenon of seabird attraction to lights. We believe the most urgently needed actions to mitigate and understand light-induced mortality of seabirds are estimation of mortality and effects on populations; determination of threshold light levels and safe distances from light sources; documentation of the fate of rescued birds; improvement of rescue campaigns, particularly in terms of increasing recovery rates and level of care; and research on seabird-friendly lights to reduce attraction.

Characterisation of functional domains in fission yeast Ams2 that are required for core histone gene transcription.

Histone gene expression is regulated in a cell cycle-dependent manner, with a peak at S phase, which is crucial for cell division and genome integrity. However, the detailed mechanisms by which expression of histone genes are tightly regulated remain largely unknown. Fission yeast Ams2, a GATA-type zinc finger motif-containing factor, is required for activation of S phase-specific core histone gene transcription. Here we report the molecular characterisation of Ams2. We show that the zinc finger motif in Ams2 is necessary to bind the histone gene promoter region and to activate histone gene transcription. An N-terminal region of Ams2 acts as a self-interaction domain. Intriguingly, N-terminally truncated Ams2 binds to the histone gene promoters, but does not fully activate histone gene transcription. These observations imply that Ams2 self-interactions are required for efficient core histone gene transcription. Moreover, we show that Ams2 interacts with Teb1, which itself binds to the core histone gene promoters. We discuss the relationships between Ams2 domains and efficient transcription of the core histone genes in fission yeast.

High levels of isotope elimination improve precision and allow individual-based measurements of metabolic rates in animals using the doubly labeled water method.

Doubly labeled water (DLW) can be used to measure energy expenditure in free-ranging animals, but questions have been raised about its accuracy in different species or contexts. We investigated whether differences in the extent of isotope elimination affects the precision and accuracy of the DLW method, which can vary according to the experimental design or metabolic rate of the species. Estimated total energy expenditure by the DLW method (TEEdlw) was compared with actual total energy expenditure simultaneously measured via respirometry (TEEresp) in streaked shearwaters Calonectris leucomelas, a pelagic seabird. Subjects were divided into three groups with different experimental conditions: at rest on the ground for 24 h (Group A) or for 48 h (Group B), and at rest on the water for 24 h (Group C). TEEdlw in Group A matched TEEresp, whereas there was an overestimation of TEEdlw in both Groups B and C compared with TEEresp. However, compared with Group A, TEEdlw in Groups B and C had reduced the isotopic analytical variability and thus higher precision. The best regression model (TEEdlw = 1.37 TEEresp - 14.12) showed a high correlation (R(2) = 0.82) between TEEdlw and TEEresp and allows a correction factor for field metabolic rates in streaked shearwaters. Our results demonstrate that the commonly made assumption that the DLW method is not appropriate for individual-based estimates may be incorrect in certain circumstances. Although a correction factor may be necessary when using the DLW method to estimate metabolic rate, greater levels of isotope eliminations provides DLW estimates with high precision, which can adequately represent relative individual estimates. Nevertheless, the DLW method, should be used with caution when characterizing interspecies difference of energy expenditures.

Applicability of the doubly labelled water method to the rhinoceros auklet, Cerorhinca monocerata.

The doubly labelled water (DLW) method is an isotope-based technique that is used to measure the metabolic rates of free-living animals. We validated the DLW method for measuring metabolic rates in five rhinoceros auklets (Cerorhinca monocerata) compared with simultaneous measurements using the respirometric method. We calculated the CO(2) production rate of four auklets (mean initial body mass: 552 g±36 s.d.) injected with DLW, using the one- and two-pool models. The metabolic rate during the 24-h measurements in a respirometric chamber for resting auklets averaged 16.30±1.66 kJ h(-1) (n = 4). The metabolic rates determined using the one- and two-pool models in the DLW method for the same period as the respirometric measurement averaged 16.61±2.13 kJ h(-1) (n = 4) and 16.16±2.10 kJ h(-1) (n = 4), respectively. The mean absolute percent error between the DLW and respirometric methods was 8.04% using the one-pool model and was slightly better than that with the two-pool model. The differences in value between the DLW and respirometric methods are probably due to oxygen isotope turnover, which eliminated only 10-14% of the initial enrichment excess.