Many genetic disorders are caused by faulty versions of a single gene. In the last decade, scientists have made tremendous strides in correcting these faults through “gene therapy”—using viruses to sneak in working versions of the affected genes.
But some disorders pose greater challenges. Down’s syndrome, for example, happens when people are born with three copies of the 21st chromosome, rather than the usual two. This condition, called trisomy, leads to hundreds of abnormally active genes rather than just one. You cannot address it by correcting a single gene. You’d need a way of shutting down an entire chromosome.
But half of us do that already. Women are masters of chromosomal silencing.
Women are born with two copies of the X chromosome, while men have just one. This double dose of X-linked genes might cause problems, so women inactivate one copy of X in each cell.
This is the work of a gene called XIST (pronounced “exist”). It produces a large piece of RNA (a molecule closely related to DNA) that coats one of the two X chromosomes and condenses it into a dense, inaccessible bundle. It’s like crunching up a book’s pages to make them unreadable and useless. XIST exists on the X chromosome, so that’s what it silences. But it should be able to shut down other chromosomes too, if we could just insert it into the right place.
That’s exactly what Jun Jiang from the University of Massachusetts Medical School has done: she used XIST to shut down chromosome 21. “Most genetic diseases are caused by one gene, and gene therapies correct that gene,” says Jeanne Lawrence, who led the study. “In this case, we show that you can manipulate one gene and correct hundreds.” It’s chromosome therapy, rather than gene therapy.
So far, the team have only done this in Down’s syndrome cells, grown in a laboratory, so the technique is a very long way from any clinical use. But it’s a promising first step, and other scientists are very excited. “It’s an amazing paper,” says Elizabeth Fisher from University College London, who studies Down’s syndrome. “The fact that they have silenced the entire chromosome will really help people to dissect what’s going wrong in Down’s syndrome.”
Lawrence has spent years studying XIST, and has always thought about applying this work to Down’s syndrome. After all, she used to provide counselling for parents whose babies are born with disabilities, and she regularly talks to families who are affected by Down’s, some of whom talk at the genetics course she runs. But while using XIST to inactivate chromosome 21 was an obvious strategy, it was also a risky one.
For a start, XIST is huge—far larger than any other gene that has been deliberately inserted into a genome before. If the team got it into the right place, would it actually silence chromosome 21 without killing the cell? And if it worked, what would stop it from silencing all three copies rather than just one? “None of these challenges made the project impossible, but collectively they made it pretty improbable,” says Lawrence. “We didn’t know if we’d spend years not getting anything to work.”
And yet, after six years of toil, it worked. Jiang used enzymes called zinc finger nucleases, which cut DNA at very specific points, to smuggle the giant XIST gene into a pre-defined spot on the 21st chromosome. She did this in cells from a boy with Down’s syndrome, which had been reprogrammed into a stem cell-like state. XIST did its thing, “painting” one of the three chromosome-21s, and condensing it into a tight bundle. The genes on that copy were almost totally inactivated.
In this study, Jiang ensured that XIST only shut down one of the three chromosomes by tweaking its concentration. In the future, the team might target it to sequences found in only one of the three copies.
But does inactivating a copy of chromosome 21 achieve anything useful? Jiang saw some promising signs. For example, after XIST, the Down’s cells grew more quickly, produced larger colonies, and were far better at dividing into neuron-making cells. This supports the idea that people with Down’s syndrome can’t make enough cells (and neurons, in particular) as they grow up.
“It’s an extremely exciting development. It’s somewhat surprising that it took so long for someone to apply this to chromosome 21, but the group had to overcome some very significant technical challenges,” says Roger Reeves from Johns Hopkins University. “The next step will be to silence an extra chromosome in an animal, as opposed to a dish of cells.” For example, they could try the technique on mice that have been bred with extra copies of chromosome 21.
Even if that worked, it would be very challenging to use the XIST technique in people—you’d need to get the giant gene into the right cells at the right stage. “I doubt that XIST by itself has the potential to become a therapeutic agent in patients,” says Stylianos Antonarakis from the University of Geneva.
Lawrence agrees, but she thinks there might be exceptions. For example, many children with Down’s develop myoproliferative disease, where they produce too many blood cells and run a high risk of leukaemia. If doctors saw kids with this condition, it might be possible to activate XIST in their blood stem cells, to prevent them from developing cancer. “That’s one of the more likely possible uses,” says Lawrence.
The study also has more immediate benefits: “It’s a way of getting at the biology that underlies the different aspects of Down’s,” says Lawrence. The syndrome includes dozens of symptoms across many different organs, including intellectual disabilities, heart problems, leukaemia and Alzheimer’s at an early age. Matching these up to the hundreds of genes on chromosome 21 has been a herculean task. “There are many studies that point to different genes but it’s still a pretty confused field,” says Lawrence.
Her team’s work could help. Scientists could activate XIST in one of two groups of identical cells, and watch what happens to the rest of their genes. They could do this in neurons, heart cells, or any of the other tissues that are affected in Down’s syndrome. They could also test drugs that are designed to alleviate the syndrome’s symptoms. And, as Antonarakis says, scientists could do this not just for Down’s syndrome, but for the many other disorders that are caused by unusual number of chromosomes.
Jiang’s work also confirms something important about XIST—it evolved to shut down the X chromosome, but it works on all of them. “It must be acting on something that’s found on all chromosomes,” says Lawrence. She thinks it might recognise repetitive bits of DNA that are found throughout our genome, but have no obvious purpose.
Indeed, Lawrence suspects that her work on XIST and Down’s might eventually tell us more about how the genome is organised. XIST is one of several pieces of RNA that are transcribed from the genome, but never used to make proteins. Because of its large size, it’s classified as a “long, non-coding RNA” or lncRNA—a group that includes tens of thousands of members. A minority of these, like XIST, clearly help to control how other genes are used, but there’s a lot of debate about what the rest do, if anything (see Carl Zimmer’s post for more).
Lawrence’s team have moved beyond this debate, and are one of the first to actually use a lncRNA to target and silence a set of genes. “That’s one of the aspects that makes it so exciting,” says Mitchell Guttman from the California Institute of Technology, who studies lncRNA and recently showed how XIST finds its way around the X chromosome. “The field will surely build upon this in the future as it continues to dissect the roles of other lncRNAs and learns more about the principles governing their localization and function.”
Reference: Jiang, Jing, Cost, Chiang, Kolpa, Cotton, Carone, Carone, Shivak, Guschin, Pearl, Rebar, Byron, Gregory, Brown, Urnov, Hall & Lawrence. 2013. Translating dosage compensation to trisomy 21. Nature