Biological aspects of cytosine methylation in eukaryotic cells.

The existence in eukaryotes of a fifth base, 5-methylcytosine, and of tissue-specific methylation patterns have been known for many years, but except for a general association with inactive genes and chromatin the exact function of this DNA modification has remained elusive. The different hypotheses regarding the role of DNA methylation in regulation of gene expression, chromatin structure, development, and diseases, including cancer are summarized, and the experimental evidence for them is discussed. Structural and functional properties of the eukaryotic DNA cytosine methyltransferase are also reviewed.     

Biological aspects of cytosine methylation in eukaryotic cells

The existence in eukaryotes of a fifth base, 5-methylcytosine, and of tissue-specific methylation patterns have been known for many years, but except for a general association with inactive genes and chromatin the exact function of this DNA modification has remained elusive. The different hypotheses regarding the role of DNA methylation in regulation of gene expression, chromatin structure, development, and diseases, including cancer are summarized, and the experimental evidence for them is discussed. Structural and functional properties of the eukaryotic DNA cytosine methyltransferase are also reviewed.     

Emerging connections between DNA methylation and histone acetylation

Modifications of both DNA and chromatin can affect gene expression and lead to gene silencing. Evidence of links between DNA methylation and histone hypoacetylation is accumulating. Several proteins that specifically bind to methylated DNA are associated with complexes that include histone deacetylases (HDACs). In addition, DNA methyltransferases of mammals appear to interact with HDACs. Experiments with animal cells have shown that HDACs are responsible for part of the repressive effect of DNA methylation. Evidence was found in Neurospora that protein acetylation can in some cases affect DNA methylation. The available data suggest that the roles of DNA methylation and histone hypoacetylation, and their relationship with each other, can vary, even within an organism. Some open questions in this emerging field that should be answered in the near future are discussed.     

Geminivirus DNA replication

Geminiviruses are DNA viruses which infect plants. They have a small genome and encode only a few proteins. Therefore, their DNA replication cycle relies largely on the use of cellular DNA replication proteins. The strategy used by geminiviruses to replicate their single-stranded DNA (ssDNA) genome consists of a first stage of conversion of ssDNA into double-stranded DNA (dsDNA) intermediates and, then, the use of dsDNA as a template to amplify viral dsDNA and to produce mature ssDNA genomes by a rolling-circle replication mechanism. In addition, the accumulating evidence indicates that viral DNA replication is somehow coupled to the cell cycle regulatory network of the infected cell. For these reasons, geminiviruses are excellent model systems to understand the regulation of DNA replication and cell cycle in plant cells. Recent years have witnessed significant progress in the identification of cis-acting signals and their interaction with trans-acting factors that contribute to geminivirus origin function. These and other aspects of the geminivirus DNA replication cycle will be reviewed.     

Hsp90 binds CpG oligonucleotides directly: implications for hsp90 as a missing link in CpG signaling and recognition

CpG motifs originating from bacterial DNA (CpG DNA) can act as danger signals for the mammalian immune system. These CpG DNA motifs like many other pathogen-associated molecular patterns are believed to be recognized by a member of the toll-like receptor family, TLR-9. Here we show results suggesting that heat shock protein 90 (hsp90) is also implicated in the recognition of CpG DNA. Hsp90 was characterized as a binder to oligodeoxynucleotides (ODNs) containing CpG motifs (CpG ODNs) after several purification steps from crude protein extracts of peripheral blood mononuclear cells. This finding was further supported by direct binding of CpG ODNs to commercially available human hsp90. Additionally, immunohistochemistry studies showed redistribution of hsp90 upon CpG ODN uptake. Thus, we propose that hsp90 can act as a ligand transfer molecule and/or play a central role in the signaling cascade induced by CpG DNA. Received 18 December 2002; accepted 6 January 2002 RID="*" ID="*"Corresponding author. B. Agerberth and G. H. Gudmundsson contributed equally to this work.     

Epigenetic mechanisms in mammals

DNA and histone methylation are linked and subjected to mitotic inheritance in mammals. Yet how methylation is propagated and maintained between successive cell divisions is not fully understood. A series of enzyme families that can add methylation marks to cytosine nucleobases, and lysine and arginine amino acid residues has been discovered. Apart from methyltransferases, there are also histone modification enzymes and accessory proteins, which can facilitate and/or target epigenetic marks. Several lysine and arginine demethylases have been discovered recently, and the presence of an active DNA demethylase is speculated in mammalian cells. A mammalian methyl DNA binding protein MBD2 and de novo DNA methyltransferase DNMT3A and DNMT3B are shown experimentally to possess DNA demethylase activity. Thus, complex mammalian epigenetic mechanisms appear to be dynamic yet reversible along with a well-choreographed set of events that take place during mammalian development.     

The karyotype and genome structure of the pirate perchAphredoderus sayanus (Aphredoderidae: Teleostei)

Summary The standard karyotype, genome size (DNA content), and genomic DNA base composition and distribution of the relict paracanthopterygian fish,Aphredoderus sayanus, were investigated. Several features of theA. sayanus genome appear to be derived rather than primitive conditions. These include a large number (at least 10 pairs) of bi-armed chromosomes, a low genome size, and high DNA asymmetry. This may indicate thatA. sayanus is not a typical paracanthopterygian fish in terms of its genome structure.     

Ultrastructural localization of 5-methylcytosine on DNA and RNA

DNA methylation is the major epigenetic modification and it is involved in the negative regulation of gene expression. Its alteration can lead to neoplastic transformation. Several biomolecular approaches are nowadays used to study this modification on DNA, but also on RNA molecules, which are known to play a role in different biological processes. RNA methylation is one of the most common RNA modifications and 5-methylcytosine presence has recently been suggested in mRNA. However, an analysis of nucleic acid methylation at electron microscope is still lacking. Therefore, we visualized DNA methylation status and RNA methylation sites in the interphase nucleus of HeLa cells and rat hepatocytes by ultrastructural immunocytochemistry and cytochemical staining. This approach represents an efficient alternative to study nucleic acid methylation. In particular, this ultrastructural method makes the visualization of this epigenetic modification on a single RNA molecule possible, thus overcoming the technical limitations for a (pre-)mRNA methylation analysis.     

DNA hypomethylation in the origin and pathogenesis of human diseases

The pathogenesis of any given human disease is a complex multifactorial process characterized by many biologically significant and interdependent alterations. One of these changes, specific to a wide range of human pathologies, is DNA hypomethylation. DNA hypomethylation signifies one of the major DNA methylation states that refers to a relative decrease from the “normal” methylation level. It is clear that disease by itself can induce hypomethylation of DNA; however, a decrease in DNA methylation can also have an impact on the predisposition to pathological states and disease development. This review presents evidence suggesting the involvement of DNA hypomethylation in the pathogenesis of several major human pathologies, including cancer, atherosclerosis, Alzheimer’s disease, and psychiatric disorders. The views expressed in this paper do not necessarily represent those of the US Food and Drug Administration.