5-hydroxymethylcytosine’s (5-hmC) discovery last year in mammalian DNA (Kriaucionis & Heintz, Science, 2009. and Tahiliani et al., Science, 2009), left the epigenetics community more shaken than a 007 martini. Although its broader role (if any) in epigenetics remains a mystery today, early evidence suggests a few putative mechanisms that could have big implications.
Last year’s discovery of 5-hydroxymethylcytosine (5-hmC) caused some epigenetics researchers to suffer a crisis of confidence, provoking many hours of soul-searching questions such as “are my methylation data really accurate?” and “what if what I thought was gene silencing is really activation?” Although current estimates place 5-hmC levels at only a fraction of the 5-mC content in most cells, it could still be enough to foul up your data interpretation, especially if 5-hmC’s function ends up being antithetical to 5-mC’s as some researchers have proposed.
One of the great things about science is that whenever a new problem presents itself, there are always some members of the community who set out to find innovative solutions. In this case, it’s been a bear to detect and quantify 5-hydroxymethylcytosine (5-hmC) but some cagey researchers at the Ludwig Maximilians University (LMU) in Munich, Germany, have established a simple and accurate method to quantify global levels of genomic 5-hmC.
To say researchers have been pretty amped about 5-Hydroxymethylcytosine (5-hmC) this last year would be an understatement. Is this 6th base for real, or is it some kind of intermediate of it’s more abundant cousin 5-mC? So far its low key presence has been detected in embryonic stem cells, brain, and other organs but there are still far more questions than answers for 5-hmC.
Small pores aren’t just highly sought after in the cosmetics industry. They can really lend a hand in smoothing out the rougher epigenetics experimental challenges researchers face nowadays. Just ask Dr. Meni Wanunu and Marija Drndic at the University of Pennsylvania, along with colleagues from New England Biolabs, Boston University, and Temple University, who put solid-state nanopores to work recently and obtained some glowing DNA methylation profiling results.
A UK team used MeDIP-Seq and hMeDIP-Seq to look at genome-wide patterns of 5mC and 5hmC in mouse ES cells and differentiating embryoid bodies. They found interesting differences in 5mC/5hmC distributions among genomic regions and cell types:
A clever team of researchers has developed two independent approaches for the genome-wide mapping of 5hmC, neither of which relies on 5hmC antibodies.
The team from New England Biolabs and UCLA kicked off their project by developing a method to detect and measure 5-hmC.
Things got even more interesting earlier this year when 5-formylcytosine(5fC) and 5-carboxylcytosine (5caC) was found to be a part of that same pathway. Now, some recent work out Yi Zhang’s lab at the University of North Carolina suggests that 5fC and 5caC appear to be more than just intermediates.
With this and several other methods, they generated detailed genome-wide epigenetic maps of 5-hmC in the mouse cerebellum and hippocampus seven days after birth, at six weeks old, and at one year old leading to lots of new insights into 5-hmC.
Dr. Wolf Reik discusses the challenges in DNA methylation sequencing and the next big swings in epigenetic modification sequencing and discovery.
Scientists from the Babraham Institute and University of Cambridge solved the 5hmC sequencing problem by implementing a new twist on the familiar bisulfite conversion method.
A talented team of scientists from Taiwan recently showed that two members of the DNA methyltransferase (DNMT) family, DNMT3a and DNMT3b are more than just methyltransferases; they fill a role as dehydroxymethylases as well.
When slick, new methods surface that tackle some tough challenges like single nucleotide resolution 5-hmC profiling, researchers’ excitement is quickly tempered by the experimental costs required to execute the latest and greatest technique. A nice workaround to this issue can be achieved by combining 5-hmC enrichment sequencing with restriction digestion-based targeted validation.
A couple of years ago, researchers keen to analyze 5-hmC had a couple of options, but those early approaches couldn’t quite satisfy all of their increasing demands. Fast-forward a bit and it’s a very different story. Today there’s a plethora of options, each with its own strengths and weaknesses. As a result, one of the biggest challenges researchers currently face in 5-hmC analysis arises long before experiments are conducted: choosing the right tool for the job.
Called “hydroxymethylation and methylation sensitive tag sequencing” (HMST-Seq), the new approach is cost-effective and high-res.
A talented crew of researchers led by Dr. Yuval Ebenstein (Tel-Aviv University, Israel) recently pioneered a technique that gives anyone the ability to assay 5hmc “nanodrop” style. In their paper, they presented proof-of-principle experimentation. They labeled DNA and then sandwiched it between two glass slides to stretch it.. The DNA was then imaged on a fluorescent microscope and 5hmC sites were counted as red spots along the DNA. This gave the exact 5hmC to nucleotide ratio in the sample.
We'd love to hear from you. Please send questions or feedback to the below email addresses.
Before contacting us, you may wish to visit our FAQs page which has lots of useful info on Tiki-Toki.
We can be contacted by email at: hello@tiki-toki.com.
You can also follow us on twitter at twitter.com/tiki_toki.
If you are having any problems with Tiki-Toki, please contact us as at: help@tiki-toki.com
Close