On Tuesday, the team behind the plan to bring mammoth-like animals back to the tundra announced the creation of what it is calling wooly mice, which have long fur reminiscent of the woolly mammoth. The long fur was created through the simultaneous editing of as many as seven genes, all with a known connection to hair growth, color, and/or texture.
But don’t think that this is a sort of mouse-mammoth hybrid. Most of the genetic changes were first identified in mice, not mammoths. So, the focus is on the fact that the team could do simultaneous editing of multiple genes—something that they’ll need to be able to do to get a considerable number of mammoth-like changes into the elephant genome.
Of mice and mammoths
The team at Colossal Biosciences has started a number of de-extinction projects, including the dodo and thylacine, but its flagship project is the mammoth. In all of these cases, the plan is to take stem cells from a closely related species that has not gone extinct, and edit a series of changes based on the corresponding genomes of the deceased species. In the case of the mammoth, that means the elephant.
But the elephant poses a large number of challenges, as the draft paper that describes the new mice acknowledges. “The 22-month gestation period of elephants and their extended reproductive timeline make rapid experimental assessment impractical,” the researchers acknowledge. “Further, ethical considerations regarding the experimental manipulation of elephants, an endangered species with complex social structures and high cognitive capabilities, necessitate alternative approaches for functional testing.”
So, they turned to a species that has been used for genetic experiments for over a century: the mouse. We can do all sorts of genetic manipulations in mice, and have ways of using embryonic stem cells to get those manipulations passed on to a new generation of mice.
For testing purposes, the mouse also has a very significant advantage: mutations that change its fur are easy to spot. Over the century-plus that we’ve been using mice for research, people have noticed and observed a huge variety of mutations that affect their fur, altering color, texture, and length. In many of these cases, the changes in the DNA that cause these changes have been identified.
That gets into the nature of this paper. It’s possible to use the mice to test the mutations that are based on the differences between the mammoth and elephant genomes, selecting a few that we might expect to alter fur. For this publication at least, however, the team chose to focus largely (though not exclusively) on genetic changes that have been identified in mice. That places the emphasis on the gene editing.
Or at least the scientific emphasis. The company is still announcing them as the “Colossal Woolly Mouse,” even though the connection to the woolly mammoth is relatively minimal.
The editorial pipeline
Still, Colossal has developed an impressive editing capacity. The team, which included a number of academic researchers, used variations of two different approaches to gene editing. One of these is based on the CRISPR/Cas9, which makes a break in both strands of DNA’s double helix at a specific sequence that matches what’s called a guide RNA.
In the second set of experiments, the researchers used the targeting and guide RNAs of the CRISPR/Cas system, but linked them to a protein called a cytosine base editor. This protein essentially lops a small part of the DNA base cytosine, or C, converting it into a thymidine, or T. That can introduce smaller and more specific mutations than CRISPR/Cas.
In both cases, the researchers simply mixed the proteins and guide RNA needed, and then used rapidly shifting electrical currents to open the membranes of mouse stem cells, allowing the complexes to diffuse into the cell and engage in DNA editing.
The researchers went through a round of testing different guide RNAs to each of the targeted genes to identify ones that could trigger specific editing. Then, using those, they tested both systems in stem cells. In general, the CRISPR/Cas9 system created more cuts in the DNA. That meant more of the target genes were edited, and it was more likely that the copy of each gene on both of the sets of chromosomes was edited at the same time. But it also meant the system was more likely to create off-target effects, editing parts of the genome distant from the one that the guide RNA targeted.
In contrast, the system that used the cytosine base editor was far less active, to the extent that the researchers had to devise a system to identify the cells where no editing whatsoever took place so they could be excluded from any further work. In addition, the cytosine base editor tended to edit fewer of the sites, and was more likely to edit just one of the two chromosomes. But, on the plus side, there were far fewer off-target edits.
Getting woolly
The researchers selected a large variety of genes that affect mouse fur, from developmental regulators that alter the behavior of the follicle cells to the keratin genes that encode the primary structural component of the fur itself. Depending on exactly how many of the genes were edited—as many as seven were edited in some experiments—the team saw different fur phenotypes. As the number of altered genes went up, however, the mice ended up with a long, golden coat, in some cases with a shaggier look due to kinked hair shafts.
Separately, they also tested a mutation in a gene that alters fat metabolism, modeled after a change found in the mammoth genome. But that didn’t produce any obvious changes.
But, as noted above, most of these changes mimic things that have been seen in mice in the past. The real story is about the advancements in gene editing that allow the researchers to target multiple genes at once with relatively high efficiency. It’s a nice technical achievement.
But it’s still well short of what they’ll want to be able to do eventually. The changes they targeted mostly result in truncated, non-functional versions of the protein these genes encode, which are relatively easy to make via gene editing. And, for the mammoth project, they’re ultimately going to want to make dozens to over a hundred changes. It’s unclear whether they can simply keep boosting the number of genes they try to edit in a single go, or if this might ultimately require multiple rounds of editing in stem cells before producing any embryos.
So, overall, the news seems to have a bit of everything: marketing hype, real technological progress, and challenges that have yet to be resolved.