Epigenetic studies add an often critical dimension to the understanding of biology. With implications ranging from gene regulation to pathogen virulence, epigenetic changes play an important role in shaping phenotypes and affecting disease. In recent years, it has become clear that the impact of epigenetics is more common and complex than previously suspected, underscoring the need to conduct in-depth studies of these mechanisms.
Despite the widespread recognition of the need to study epigenetics, technology has been a limiting factor in the ability to detect and identify these DNA base modifications. Bisulfite sequencing is a frequently used approach to detect 5-methylcytosine (5mC) marks, but it relies on a cytosine conversion method that can be incomplete, resulting in false positive identifications. It also can have trouble distinguishing between 5mC and other forms of methylation. Another method for studying epigenetics involves microarrays, which are additionally limited to querying known methylation sites and thus cannot detect novel sites.
Recently, scientists have begun to successfully use single molecule, real-time (SMRT) sequencing technology to analyze epigenetics. Unlike other sequencing methods, epigenetic marks are preserved in this approach due to the absence of DNA amplification; as the sequencer moves along the DNA strand, epigenetic data is recorded together with nucleic acid sequences. Scientists have used this method to analyze whole epigenomes of microbes, which may ultimately provide critical insight for drug screening and microbial targeting. Here, is highlighted important new studies of epigenomes and how these discoveries could inform new approaches for drug discovery and development research.
In a study recently published in PLoS Genetics from scientists at the Joint Genome Institute and other organizations, scientists conducted a large-scale survey of genome-wide methylation in more than 200 bacterial and archaeal species (“The Epigenomic Landscape of Prokaryotes”). Their results demonstrate that epigenetic marks across bacteria were far more common than expected, with some form of methylation found in 93 percent of organisms analyzed. The team studied several types of methylation, including 5mC as well as 6-methyladenine (6mA) and 4-methylcytosine (m4C).
Through the project, scientists identified hundreds of methylated motifs, associated responsible methyltransferases with their methylation specificities, and found numerous novel restriction-modification systems. Nearly half of the species studied had orphan methyltransferases, indicating that these organisms use methylation for genome regulation in addition to its more established role in genome protection. The team hypothesized that such extensive methylation in these organisms likely indicates that the epigenome has many more functions than those currently recognized.
In-depth studies of individual organisms have shed light on a phase-variation-based epigenetic mechanism — known as a phasevarion — that can alter pathogenicity in bacteria. A recent publication from scientists at the University of Leicester and other institutes reported a restriction-modification system that acts as a genetic switch in Streptococcus pneumoniae, randomly assigning cells to one of six biological phases that range from benign to highly pathogenic (“A random six-phase switch regulates pneumococcal virulence via global epigenetic changes”). This methylation-dependent switch essentially requires an infected host to combat multiple different bacteria at once, enhancing the pathogen’s ability to survive attack from the immune system. Other recent studies have brought similar mechanisms to light, finding that phasevarions play critical roles in regulating virulence in many bacteria commonly responsible for important diseases (for example: “A biphasic epigenetic switch controls immunoevasion, virulence and niche adaptation in non-typeable Haemophilus influenzae” and “Specificity of the ModA11, ModA12 and ModD1 epigenetic regulator N6-adenine DNA methyltransferases of Neisseria meningitides”).
This enhanced understanding of how microbes use epigenetics, and particularly the role of phasevarions in pathogenicity, provides intriguing new evidence that may be critical for the discovery and development of new drugs or vaccines targeting these organisms.
Novel methylation patterns
Other research teams have used SMRT sequencing technology to analyze methylation in previously inaccessible samples or in organisms not known to use epigenetic mechanisms.
A recent report of genome-wide methylation analyses for bacteria found in human microbiome samples is thought to be the first application of this approach to such samples (“The methylome of the gut microbiome: disparate Dam methylation patterns in intestinal Bacteroides dorei”). In it, scientists from the University of Florida and other institutions studied bacteria isolated from fecal samples and found highly divergent methylation profiles, suggesting that bacterial function in these communities cannot be inferred just from DNA sequence. The findings also highlight a potential link to diabetes autoimmunity, offering new clues to understanding a complex disease.
Another project analyzed C. elegans, which was believed not to use DNA methylation, and found the first evidence of 6mA in the genome (“DNA Methylation on N6-Adenine in C. elegans”). Harvard Medical School researchers found that 0.7 percent of adenines in the worm’s genome were methylated and conducted follow-up experiments to show that these modifications were heritable, with increasing effect over generations. The findings indicate that eukaryotes may use more types of methylation than previously expected.
Throughout the genomics community, there is growing evidence that a significant amount of DNA methylation has gone unappreciated and is only now being revealed. From complex gene expression patterns regulated by phasevarions to methylation where there was thought to be none, it has become clear that epigenetic mechanisms are sophisticated, widespread, and used in unexpected ways.
Perhaps most exciting, these new studies suggest that epigenetics may have immense value for drug discovery research, where epigenetic modifications are already being evaluated as important new targets. Mounting evidence shows that methylation patterns and phasevarions may be critical regulators of pathogenicity, and that finding ways to influence these mechanisms could help shift dangerous bacteria to a more benign state. Perhaps in the future, treatment will aim not to eliminate bacteria from a host entirely but instead to convert newly acquired strains to harmless passersby.
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