Land Of The Houyhnhnms: How NGS Turned The Genomic Clock Back 700,000 Years

Some 700,000 years ago, a male horse died alone in the Canadian Arctic.  Though he met his demise during a relatively warm period for the Yukon Territory, his corpse settled between thick layers of permafrost and his bones slowly fossilized until, ten years ago, a team of highly intelligent primates unearthed one of his hoof bones.

Recognition…at last

Today, that fossil is bringing this equine posthumous recognition as the oldest creature to have its genome completely sequenced. The announcement, in the form of a recent article in Nature, unleashes a torrent of data relevant not only to equine evolution, but also paleogenomics and the limitations of sequencing ancient genomes.

Yukon Gold

Thanks to its coffin of permafrost, molecules within the fossilized bone fragment fared fairly well in the estimated 732,000 years since the horse’s foundering.  Protein mass spectrometry revealed blood and collagen peptides. Encouraged by the initial MS results, a team of European, Canadian, Chinese, Saudi and American scientists probed further, using Illumina and Helicos platforms to attempt genome sequencing of the archaic specimen. Both platforms overwhelmingly yielded contaminants (over 95%), though they obtained sufficient horse sequence (over 12 billion reads) to assemble this ancient equine genome at 1.12x coverage across the modern horse genome.

Horsey truths

One might be tempted to declare victory and move on after such an accomplishment.  After all, the previous record for ancient genome sequence (a Denisovan human) was 80,000 years. However, to verify the old horse genome’s authenticity (and to answer some long-standing questions regarding horse diversity, evolution, and domestication), the authors compared the genome of this Yukon horse with the genomes of five domesticated horses, Przewalski’s horse (the last remaining wild horse from the Mongolian steppes), a donkey, and another ‘ancient’ horse genome (this one dating back some 43,000 years). Their comparisons of the 700-millenia-old Yukon horse with the genomes of these more recent equines confirm some horse truths whilst turning others on their horsey heads!

A horse is a horse, of course, of course

The Yukon genome confirms the monophyletic nature of modern horses and their closest relatives (all surviving Equus species, apart from zebras). But based on sequence divergence between this ancient genome and more modern counterparts, we must now push the origins of today’s equines back four to four and a half million years ago, doubling paleontological estimates of merely two million years. In addition, Przewalski’s horse is truly Earth’s last remaining wild horse population, showing no signs of admixture with domestic varieties and retaining a good deal of genetic diversity despite their recent bottleneck of just 14 breeding individuals.

No grass, no horse

The ancient Yukon genome also permitted using a pairwise sequential Markovian coalescent approach to model fluctuations in horse population over hundreds of thousands of years. In general, the glaciers were not kind to our equine friends.  Periods of glacial maxima coincided with the loss of their beloved grasslands, constricting horse populations across the Northern Hemisphere. The last ice age obliterated them entirely from their native North America, leaving their reintroduction in the New World to Spanish colonists in the 16th century.

Until the cows come home

This ancient horse genome will keep equine evolutionary biologists and geneticists busy until the cows come home (pun intended). Conversely, the implications of this most archaic of genomes surpasses the interests of our hooved companions. These findings verify a crucial theory of paleogenomics, albeit with limits. It turns out that we can indeed sequence genomes from very old specimens- up to a point, and under specific circumstances.

When we die

When we die, the enzymes which protect our genomes finally stop working. This leaves DNA vulnerable to cleavage by scavengers and our own catabolic proteins. At warm temperatures, the carnage is fast and absolute- slicing and dicing genomes to their obliteration. However, in the far North, encased in Canadian permafrost, the destructive process slowed sufficiently for at least one ‘mildly’ ancient horse genome to stick around in larger pieces, until Illumina and Helicos platforms could unearth its secrets. This Yukon horse genome confirms that 700,000 years genomes are salvageable thanks to cold-preserving. Some say the upper limit can be as high as one million years, under the right (frigid) circumstances.

Gulliver’s Travels

In the final section of Gulliver’s Travels, author Jonathan Swift sends our eponymous hero to a remote land where primitive, ignorant humans serve as the beasts of burden to an intelligent and enlightened race of horses (who call themselves “Houyhnhnms”). Later, returning to his native England, poor Gulliver ignores his human companions and wastes time trying in vain to speak to any horse he encounters.  For paleogenomicists seeking to reach (or shatter) the one million year ice ceiling, the lesson here is simple- do not waste valuable time on fossils found between the Arctic and Antarctic Circles. Head to the poles.  Wrench fossils from permafrost and ice sheets and sequence the (barely) preserved DNA within. Climate change adds uncertainty, as specimens could thaw and degrade faster than we can discover them. So get going, and don’t stop to talk to every horse you meet.

Further Reading:

Orlando et al.  2013.  “Recalibrating Equus evolution using the genome sequence of an early Middle Pleistocene horse.”  Nature499: 74-81.

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