Mouse retrotransposons: sequence structure, evolutionary age, genomic distribution and function.

Retrotransposons are transposable elements that are transposed via transcription and reverse transcription. Their copies have accumulated in the genome of mammals, occupying approximately 40% of mammalian genomic mass. These copies are often involved in numerous phenomena, such as chromatin spatial organization, gene expression, development and disease, and have been recognized as a driving force in evolution. Different organisms have gained specific retrotransposon subfamilies and retrotransposed copies, such as hundreds of Mus-specific subfamilies with diverse sequences and genomic locations. Despite this complexity, basic information is still necessary for present-day genomic and epigenomic studies. Herein, we describe the characteristics of each subfamily of Mus-specific retrotransposons in terms of sequence structure, phylogenetic relationships, evolutionary age, and preference for A or B compartments of chromatin.

Sequence divergence and retrotransposon insertion underlie interspecific epigenetic differences in primates.

Changes in the epigenome can affect the phenotype without the presence of changes in the genomic sequence. Given the high identity of the human and chimpanzee genome sequences, a substantial portion of their phenotypic divergence likely arises from epigenomic differences between the two species. In this study, the transcriptome and epigenome were determined for induced pluripotent stem cells (iPSCs) generated from human and chimpanzee individuals. The transcriptome and epigenomes for trimethylated histone H3 at lysine-4 (H3K4me3) and lysine-27 (H3K27me3) showed high levels of similarity between the two species. However, there were some differences in histone modifications. Although such regions, in general, did not show significant enrichment of interspecies nucleotide variations, gains in binding motifs for pluripotency-related transcription factors, especially POU5F1 and SOX2, were frequently found in species-specific H3K4me3 regions. We also revealed that species-specific insertions of retrotransposons, including the LTR5_Hs subfamily in human and a newly identified LTR5_Pt subfamily in chimpanzee, created species-specific H3K4me3 regions associated with increased expression of nearby genes. Human iPSCs have more species-specific H3K27me3 regions, resulting in more abundant bivalent domains. Only a limited number of these species-specific H3K4me3 and H3K27me3 regions overlap with species-biased enhancers in cranial neural crest cells, suggesting that differences in the epigenetic state of developmental enhancers appear late in development. Therefore, iPSCs serve as a suitable starting material for studying evolutionary changes in epigenome dynamics during development.

The Expression Dynamics of piRNAs Derived From Male Germline piRNA Clusters and Retrotransposons.

In mammals, germ cells produce a class of small regulatory RNAs called PIWI-interacting RNAs or piRNAs, which are 25-32 nucleotides in length. The profile of testicular piRNAs changes during development. The piRNAs detected in fetal testes at embryonic day 13.5 and later are called fetal piRNAs. The piRNAs detected in testes in a period where germ cells do not yet enter the pachytene stage of meiotic prophase I are called pre-pachytene piRNAs, whereas those in testes at later postnatal days are called pachytene piRNAs. Here, to elucidate the exact expression dynamics of these piRNAs during development, we compared piRNAs present in male germ cells at different stages, which were purified by fluorescence-activated cell sorting, and those in embryonic testes. The analysis identified three distinct groups of piRNA clusters: prospermatogonial, early, and late clusters. piRNA length was largely correlated with the repertoire of PIWI-like proteins in respective germ cells; however, the late piRNA clusters tended to generate longer (PIWIL1-type) piRNAs, whereas the early clusters tended to generate shorter (PIWIL2-type) piRNAs, suggesting a cluster- or sequence-dependent mechanism for loading onto PIWI-like proteins. Retrotransposon-derived piRNAs, particularly evolutionary young retrotransposons, were abundantly produced in prospermatogonia, however, their abundance declined as development proceeded. Thus, in later stages, retrotransposon-derived piRNAs were not enriched with those from evolutionary young elements. The results revealed that, depending on the piRNA clusters from which they are derived, longer PIWIL1-type piRNAs are produced earlier, and shorter PIWIL2-type piRNAs remain in a longer period, than previously thought.

B2 SINE Copies Serve as a Transposable Boundary of DNA Methylation and Histone Modifications in the Mouse.

More than one million copies of short interspersed elements (SINEs), a class of retrotransposons, are present in the mammalian genomes, particularly within gene-rich genomic regions. Evidence has accumulated that ancient SINE sequences have acquired new binding sites for transcription factors (TFs) through multiple mutations following retrotransposition, and as a result have rewired the host regulatory network during the course of evolution. However, it remains unclear whether currently active SINEs contribute to the expansion of TF binding sites. To study the mobility, expression, and function of SINE copies, we first identified about 2,000 insertional polymorphisms of SINE B1 and B2 families within Mus musculus. Using a novel RNA sequencing method designated as melRNA-seq, we detected the expression of SINEs in male germ cells at both the subfamily and genomic copy levels: the vast majority of B1 RNAs originated from evolutionarily young subfamilies, whereas B2 RNAs originated from both young and old subfamilies. DNA methylation and chromatin immunoprecipitation-sequencing (ChIP-seq) analyses in liver revealed that polymorphic B2 insertions served as a boundary element inhibiting the expansion of DNA hypomethylated and histone hyperacetylated regions, and decreased the expression of neighboring genes. Moreover, genomic B2 copies were enriched at the boundary of various histone modifications, and chromatin insulator protein, CCCTC-binding factor, a well-known chromatin boundary protein, bound to >100 polymorphic and >10,000 non-polymorphic B2 insertions. These results suggest that the currently active B2 copies are mobile boundary elements that can modulate chromatin modifications and gene expression, and are likely involved in epigenomic and phenotypic diversification of the mouse species.

[Apoptosis-inducing properties of ent-kaurene-type diterpenoids from the liverwort Jungermannia truncata].

Ent-11alpha-hydroxy-16-kauren-15-one (1) induced apoptosis in a human leukemia cell line (HL-60 cells), however, the apoptosis-inducing properties of 1 and its related compounds remain to be proved. We examined the involvement of caspases, a family of cysteine aspartic proteases, which play a central role in induction of apoptosis, in apoptosis induced by the compounds in HL-60 cells. Treatment of the cells with compounds 1, 2 and 3 with the enone group at C-15/C-16 caused DNA fragmentation, a sign of induction of apoptosis, and proteolysis of poly(ADP-ribose) polymerase (PARP), a hallmark of caspase activation. Z-Asp-CH2-DCB, abroad spectrum inhibitor of caspases, abolished the appearance of DNA fragmentation and also significantly attenuated the cytotoxic effects. These data suggest that induction of apoptosis by 1 and some of its related compounds are dependent on caspases activation and might be partly involved in the cytotoxicity in HL-60 cells.

Mercuric chloride induces apoptosis via a mitochondrial-dependent pathway in human leukemia cells.

Mercurial compounds modulate immunologic functions by inducing cytotoxicity. Although mercury chloride (HgCl(2)) is known to induce apoptosis in various immune system cells, the mechanism of the induction of apoptosis is poorly understood. In this study, we examined the activation of caspase-3, an important cysteine aspartic protease, during HgCl(2)-induced apoptosis in a human leukemia cell line (HL-60 cells). Both DNA fragmentation, a characteristic of apoptotic cells, and proteolysis of poly(ADP-ribose) polymerase (PARP), a substrate of caspase-3, occurred at 6 h after HgCl(2) treatment in HL-60 cells. These results suggest that the activation of caspase-3 was involved in HgCl(2)-induced apoptosis. The release of cytochrome c (Cyt c) from mitochondria into the cytosol, which is an initiator of the activation of caspase cascades, was also observed in HgCl(2)-treated HL-60 cells. Moreover, the release of Cyt c from mitochondria was observed in HgCl(2)-treated mitochondria isolated from mice liver, and this was followed by mitochondrial permeability transition (PT). The PT was inhibited by cyclosporin A (CsA), a potent inhibitor of PT. CsA also suppressed the occurrence of DNA fragmentation induced by HgCl(2) treatment in HL-60 cells. Taken together, these findings indicate that HgCl(2) is a potent inducer of apoptosis via Cyt c release from the mitochondria in HL-60 cells.

Heart rate variability and arterial blood pressure variability show different characteristic changes during hemorrhage in isoflurane-anesthetized, mechanically ventilated dogs.

UNLABELLED: We assessed the changes in heart rate variability (HRV) and blood pressure variability (BPV) as indices of autonomic nervous system and volume status during hemorrhage in isoflurane-anesthetized, mechanically ventilated dogs. Nine dogs were used. They were sequentially subjected to withdrawal of 30% estimated blood volume and graded isoflurane inhalation of 1% and 2% followed by discontinuation of isoflurane and retransfusion. The power spectra of HRV and BPV were computed using the fast Fourier transformation, and were quantified by determining the areas of the spectrum in two component widths: low-frequency component (LF) (0.04-0.15 Hz) and high-frequency component (HF) (0.15-0.4 Hz). During hemorrhage and isoflurane anesthesia, both HRV-LF and HRV-HF were decreased and plateaued at the smaller concentration of isoflurane, whereas BPV-LF decreased concentration-dependently. BPV-HF showed a completely different response and increased significantly during 2% isoflurane. We speculate that HRV and BPV-LF would be affected by the autonomic nervous activity, whereas BPV-HF would depend on relative/absolute change in circulating blood volume. IMPLICATIONS: Power spectra of heart rate variability (HRV) and blood pressure variability (BPV) were computed using the fast Fourier transformation. The HRV and BPV showed their differential characteristics during hemorrhage, isoflurane anesthesia, and retransfusion, and would help to assess changes in autonomic nervous system and preload under mechanical ventilation.