The completion of the human genome, rather than being the huge breakthrough it was presented as, raised almost as many questions as it answered. Less than two percent of it encoded a protein, and only about five percent ended up being conserved relative to many of our fellow mammals. The rest of it seemed like a bit of a mess—damaged viruses, long stretches of repetitive sequence, and huge stretches devoid of any genes.
The ENCODE project was formed to make sense of that mess. A huge consortium of labs, ENCODE started performing a massive assay of pretty much everything to do with DNA: where proteins stuck to it, how it was packaged, how it was modified, etc. Now, the consortium is out with a massive number of papers—six alone in today's Nature, and dozens to follow in the days to come, scattered through many different journals. The results are staggering: over 1,500 different types of data, assayed across the whole genome, from 147 different cell types. It will feed researchers for years to come.
And it suggests that more of the chaos in the genome may be useful, although that suggestion comes with some big caveats.
First, the data. The researchers used a huge variety of techniques to look at just about anything that can happen to the DNA inside a cell. They took 119 proteins that bind to DNA and found every single site in the genome that they stuck to. They looked for signs of a chemical modification of DNA called methylation that can change the expression of genes nearby. The location of the proteins that structure chromosomes, called histones, were also checked, and areas where the chromosome structure is relatively accessible were pinned down. Regions of the chromosomes that are close to each other inside a cell were mapped. Every single RNA within a cell was sequenced and resequenced.
Once a single cell type, like liver cells, was checked, the authors moved on to look at another. And another. The end result was 1,640 different data sets, covering a total of nearly 150 different cell types. It's a staggering amount of work; calling ENCODE the Large Genome Collider would provide a real sense of the scale of this effort.
Functional or not?
What does it tell us about the genome? Some interesting things, although they're a bit tough to interpret. With data on this scale, we can't actually look at each piece of data on every piece of DNA. So we have to define some generic definitions of things like function: an RNA is made there, a protein binds there, etc. ENCODE chose to go extremely broad: "Operationally, we define a functional element as a discrete genome segment that encodes a defined product (for example, protein or non-coding RNA) or displays a reproducible biochemical signature (for example, protein binding, or a specific chromatin structure)." Basically, if anything came out of any of these assays regarding a base, it was considered functional.
And by that definition, most of the genome is doing something. The figure was about 80 percent in these assays, and by adding more cell types, the authors suggest we could get the number even higher.
But there are some problems with that figure. For instance, many of the non-functional viruses and mobile genetic elements will still contain the protein binding sites that used to be critical to their function. So even though these will be traditionally classified as "junk DNA," they'll be considered functional under the ENCODE definition. More generally, proteins that bind DNA are not very picky about their binding sites, so they will create some indication of function by accident; they can also lead to RNA production nearby, looping other bases into an apparent function.
That said, the study does provide some solid indications that more of the genome is doing useful stuff than many previous studies have estimated. More stringent criteria can be used—multiple proteins in close proximity, an accessible chromosome configuration, etc. These reduce the percentage of the genome that is likely to be doing something biologically useful down to about 10 percent—which is lower, but still a lot higher than the five percent that is conserved throughout mammalian evolution.
The authors mention a bit about where that additional five percent might come from. Some of it appears to be primate specific, in that it's common to primates but not found in other mammals. That hints that it might be doing something specific in our own lineage. Further evidence comes from the 1,000 genomes project, which suggests some of these sequences are doing human-specific things.
Although the 80 percent figure doesn't really fit my definition of functional, the authors do present a compelling case that more of the genome is doing something useful than we previously had evidence for.
Definitions with consequences
One of the other things the authors look at is all the human genetic differences that have been associated with diseases. Most of these haven't been located in genes, but the authors find they're enriched in the sequences that have met some definition of "functional." And, if you expand to look for things that are only near something that has been identified by ENCODE (rather than right at a location), the number goes up further still.
There are probably many cases where this involves a real and significant association between a change and human disease. But it's also probable that many of these are false positives, given the very high frequency of sites identified as giving a positive signal in at least some ENCODE assays.
In the long term, that could be a significant problem. ENCODE is mostly providing a resource for other researchers to understand the things they're studying, be it DNA binding proteins or human diseases. With 1,640 assays in 147 cell types, it will be very easy for them to search the ENCODE data and find a single assay that tells them what they'd like to see.
This danger is highlighted by the fact that the percentage of the genome flagged as functional is already at 80 percent, and the authors suggest that the number will go higher when they look at additional cell types and DNA binding proteins. If it reaches 100 percent (something that an ENCODE researcher has already suggested), then it will be very difficult to not come home with a positive result whenever you search the ENCODE genome data.
All of this means that ENCODE will be a tool that has to be used very carefully, and things that come out of it should be checked in detail. Still, it's a potentially powerful tool, since it provides so much information about so many different genes and cell types. Biologists will undoubtedly end up enjoying having it as much as they've enjoyed having the genome sequence in the first place.