Roughly 52,000 years ago, a woolly mammoth died in the Siberian tundra. As her body flash froze in the biting cold, something remarkable happened: Her DNA turned into a fossil. It wasn’t just the genetic letters that were remembered – the cold preserved their complex structure as well.
Fast forward to 2018, when an international expedition to the area found her preserved body. The team took small pieces of skin from her head and ear, her hair still intact.
From these samples, the scientists created a three-dimensional reconstruction of the woolly mammoth genome down to a nanometer. The results were published in cell today.
As in humans, mammoth DNA strands are tightly packed into chromosomes inside cells. These sophisticated structures are difficult to analyze in detail, even for humans, but they provide insight into which genes are turned on or off and how they are organized in different cell types.
Previous attempts to reconstruct ancient DNA had only tiny fragments of genetic sequences. The resulting DNA maps were incomplete, like trying to put together a puzzle with missing pieces.
Thanks to newly discovered flash-frozen DNA, this mammoth project—pun intended—is the first to assemble a huge ancient genome in 3D.
“This is a new type of fossil, and its scale exceeds the size of individual ancient DNA fragments — millions of times the sequence,” study author Erez Lieberman Aiden of Baylor College of Medicine said in a statement.
Aiden’s team worked closely with Love Dalén at the Paleogenetics Center in Sweden. In a separate study, Dalén’s team analyzed 21 genomes of Siberian mammoths and mapped how the species survived six millennia after a potentially catastrophic genetic bottleneck.
The genomes of mammoths were not so different from the genomes of today’s Asian and African elephants. They all have 28 pairs of chromosomes, and unlike most mammals, their X chromosomes twist into unique structures. Looking deeper, the team found genes that were turned on or off in the mammoth compared to its elephant cousins.
“Our analyzes reveal new biology,” Aiden’s team wrote in their paper.
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Ancient DNA is hard to find, but it offers invaluable clues about the evolutionary past. In the 1980s, scientists eager to explore genetic history showed that ancient DNA, however fragmented, could be extracted and sequenced in samples from an extinct member of the horse family and Egyptian mummies.
Thanks to modern DNA sequencing, the study of ancient DNA “has subsequently undergone a remarkable expansion,” Aiden’s team wrote. It is now possible to sequence the entire genomes of extinct humans, animals, plants and even pathogens spanning millions of years.
Making sense of the fragments is another matter. One way to decipher ancient genetic codes is to compare them to the genomes of their closest living cousins, such as mammoths and elephants. In this way, scientists can see which parts of the DNA sequence have remained unchanged and where evolution has swapped letters or small fragments.
These analyzes can link genetic changes to function, such as identifying which genes caused mammoths to be woolly. However, they cannot capture large differences at the chromosomal level. Because DNA is related to the 3D structure of a chromosome in order to function, sequencing its letters alone misses valuable information, such as when and where genes are turned on or off.
The master of chromosome puzzles
Enter Hi-C. The technique was developed in 2009 to reconstruct human genomes and detects interactions between different genetic sites inside a cell’s nucleus.
Here’s roughly how it works. DNA strands are like ribbons that wind around proteins in a structure resembling beads on a string. This arrangement brings the different parts of the DNA strand closer together in physical space. Hi-C will “blind” parts that are close together and mark the pairs. Alongside modern DNA sequencing, this technique creates a catalog of DNA fragments that interact in physical space. Similar to a 3D puzzle, scientists can put the pieces back together.
“Imagine you have a puzzle that has three billion pieces, but you don’t have a picture of the final puzzle to work from,” study author Marc A. Marti-Renom said in a press release. “Hi-C allows you to get a rough idea of that picture before you start putting the puzzle pieces together.”
But Hi-C may be impossible to use in old samples because the surviving fragments are so short that they have erased all chromosome shapes. Over time, they literally disappeared.
In the new study, the team developed a new technique called PaleoHi-C to specifically analyze ancient DNA.
The scientists immediately treated the samples in the field to reduce contamination. They created roughly 4.4 billion “pairs” of physically aligned DNA sequences—some interacting within one chromosome, others between two. Overall, they painted a 3D picture of the mammoth’s genetic material and what it looked like inside cells with nanoscale detail.
In the new reconstructions, the team identified chromosome territories — certain chromosomes are located in different regions of the nucleus — along with other peculiarities, such as loops that bring pairs of distant genomic sites into close physical proximity to alter gene expression. These patterns differ between cell types, suggesting that it is possible to see which genes are active, not just for the mammoth, but also compared to its closest living relative, the Asian elephant.
About 820 genes differed between the two, with 425 active in the mammoth but not the elephant, and a similar number inactivated in one but not the other. One inactive mammoth gene that is active in elephants has a human variant that is also switched off in the Nunavik Inuit, an indigenous people who thrive in the Arctic. The gene “may be relevant for adaptation to cold environments,” the team wrote.
Another inactive gene may explain how the woolly mammoth got its name. In humans and sheep, switching off the same gene can lead to excessive hair or wool growth.
“For the first time, we have woolly mammoth tissue in which we roughly know which genes were on and which were off,” Marti-Renom said in the report. “This is an extraordinary new type of data and the first measurement of cell-specific gene activity of genes in any ancient DNA sample.”
Crystallized DNA
How did the architecture of the mammoth genome remain so well preserved for more than 50,000 years?
Dehydration, which is often used to preserve food, may have been the key. Using Hi-C on fresh beef, beef after 96 hours sitting on the table or jerky after a year at room temperature, the jerky won with its durability. Even after being run over by a car, dunked in acid, and crushed with a shotgun (no joke), the dehydrated beef’s genomic architecture remained intact.
Dehydration may also have been part of the reason the mammoth specimen lasted so long. A chemical process called the “glass transition” is widely used to produce shelf-stable foods such as tortilla chips and instant coffee. Prevents pathogens from taking over or breaking down food. Mammoth DNA may also have been preserved in a glassy state called “chromoglass”. In other words, the sample was preserved for millennia by being freeze-dried.
It’s hard to say how long the DNA architecture can survive as chromoglass, but the authors estimate it’s probably over two million years. Whether PaleoHi-C can work on hot air-dried samples like those from ancient Egypt remains to be seen.
For mammoths, the next step is to examine gene expression patterns in other tissues and compare them to Asian elephants. In addition to building an evolutionary pipeline, the effort could also inform ongoing studies that seek to revive some version of the majestic beasts.
“These results have obvious implications for current mammoth eradication efforts,” study author Thomas Gilbert of the University of Copenhagen said in the report.
Image credit: Beth Zaiken
