Neanderthal Man

by Pbo, Svante

  The next morning it was difficult to get up to catch the taxi to the airport, and none of us spoke much until an hour or two into our flight to Moscow. By then, the bleak reality of our situation was slowly dawning on me, tainted by the dreariness and cold sweat of a serious hangover. Maybe they were already writing a paper in Berkeley on the Denisova bone. We had started writing a paper on our Denisova mtDNA results over Christmas, but it was now urgent that we finish this paper as soon as possible. Where would we submit it? The editors at Science were already impatiently waiting for our Neanderthal genome paper. To approach them about a different paper on a different topic might make them write us off as unable to finish one project, let alone two. So we decided to contact Nature. During a long layover at the airport in Moscow, I wrote an e-mail to Henry Gee, the senior editor who handles paleontology at Nature, and to Magdalena Skipper, the editor who handles genomics. I told them that we had a paper almost finished that described “what we interpret as a new hominin species based on a complete mitochondrial DNA sequence that diverged from the human line about twice as long ago as the Neanderthal mtDNA.” I was all too aware that the publication process could drag on for many months. It could even end in rejection, after months of dithering with reviewers and editors, after which we would need to submit to another journal and endure another similarly lengthy process. I didn’t want that to happen this time so I told them that we had direct competition and would be grateful if the paper could be handled quickly. An hour and fifteen minutes later, Henry Gee replied with “How exciting! Prediction is very hard, especially about the future. However, when you send it in, we’ll give it topmost priority.”

  Figure 22.3. The Denisova molar. Photo: B. Viola, MPI-EVA.

  As soon as we were back in Leipzig we finished the manuscript, which we entitled “The Complete mtDNA Genome of an Unknown Hominin from Southern Siberia,” and sent it off to Nature. It was a unique paper. For the first time ever, a new form of extinct humans was described from DNA sequence data alone, in the total absence of any skeletal remains. Given that the mtDNA was so different from that of both modern humans and Neanderthals, we felt sure that we had found a new form of extinct human. In fact, we were so taken with this idea that, after some discussion, we decided to describe it as a new species, which we called Homo altaiensis.

  However, I felt vaguely uneasy about suggesting a new species and soon had second thoughts. To me, taxonomy, the classification of living organisms into species, genera, orders, and so on, is a sterile academic exercise, particularly when discussing extinct human forms. Whenever my students send me manuscripts in which they use Linnaean Latin names for groups that are commonly known—for example, “In order to better understand the pattern of genetic variation in Pan troglodytes, we sequenced . . . ”

  —I always delete the Latin and sometimes even snidely ask who they are trying to impress by saying “Pan troglodytes” instead of “chimpanzees.” Another reason I dislike taxonomy is that it has a tendency to elicit scientific debates that have no resolution. For example, if researchers refer to Neanderthals as “Homo neanderthalensis,” they indicate that they regard them as a separate species, distinct from “Homo sapiens.” This invariably infuriates multiregionalists, who see continuity from Neanderthals to present-day Europeans. If researchers say, “Homo sapiens neanderthalensis,” they indicate that they see them as a subspecies, on par with “Homo sapiens sapiens.” This invariably infuriates proponents of the strict out-of-Africa hypothesis. These arguments I prefer to avoid, and although we had by now shown (but not yet published) that there had been mixing between Neanderthals and modern humans, I knew that taxonomic wars over Neanderthal classification would continue, since there is no definition of a species perfectly describing the case. Many would say that a species is a group of organisms that can produce fertile offspring with each other and cannot do so with members of other groups. From that perspective we had shown that Neanderthals and modern humans were the same species. However, this concept has its limitations. For example, polar bears and grizzlies can (and occasionally do) produce fertile offspring with each other when they meet in the wild. Yet polar bears and grizzlies look and behave differently, and are adapted to different lifestyles and environments. It would seem rather arbitrary, if not outright ridiculous, to regard them as one and the same species. We didn’t know whether the fact that Neanderthals contributed perhaps 2 to 4 percent of the genes of many present-day humans meant that they were the same or different species. So it was ironic that, having always refrained from using a Latin name for Neanderthals in our papers, I was now on the verge of introducing a new Linnaean species designation myself.

  Despite my misgivings about fruitless taxonomic debates, I felt I had some reasons for this digression from my principles. The mtDNA of the Denisova individual was about twice as different from the mtDNAs of modern humans as was the mtDNA of Neanderthals. That probably made them more like H. heidelbergensis, who did get to have their own Latin species name. But there was also vanity involved. Not many people get to name a new hominin species, which made it tempting to do so, even more so because this was the first time it would be done based solely on DNA data. However, the deciding argument came both from some people in our group and from Henry Gee at Nature. He pointed out that if we didn’t take the initiative and give this hominin group a species name, someone else would. And that person might come up with a name we didn’t like. So, after deliberating with Anatoly and the team who had excavated the crucial finger bone, we settled on provisionally naming it Homo altaiensis.

  Nature kept its promise to process our paper quickly: eleven days after our submission, we received comments from four anonymous reviewers. They all praised the technical aspects of the paper but they were divided on the issue of naming a new species. Two reviewers voiced concerns that we might actually have sequenced a late Homo erectus. They felt that if H. erectus had had continuous contact with groups in Africa, they may not show an mtDNA divergence as deep as their first exit out of Africa some 2 million years ago. I doubted this. But the fourth reviewer made the point that saved us from ourselves. He or she said that “once a name is in the taxonomic literature, it cannot be withdrawn later. So such provisional naming is not wise, I believe.” When I read this, I realized we had been foolish.

  In the meantime, it dawned on us that the very large amounts of mtDNA that Johannes had been able to capture from the Denisova DNA libraries meant we would be able to sequence quite a bit of this individual’s nuclear genome. This would settle its relationships both to Neanderthals and to modern humans in a definitive way as well as its possible status as a new species. We rewrote the manuscript and removed any reference to a new species. Instead, we said that “nuclear DNA sequences are needed to clarify definitively the relationship of the Denisova individual to present-day humans and Neanderthals.” We sent it back to Nature, where it appeared in early April.{63} As events would show, we had reason to be grateful that we had not named it a new species.

  Chapter 23

  A Neanderthal Relative

  ________________________________

  We began the nuclear DNA sequencing from the libraries that Johannes had prepared from the bone as soon as we could. The results were stunning. When Udo mapped them to the human genome, he found matches for about 70 percent of all DNA fragments. Yet contamination with modern human DNA, as judged from the mtDNA results, was extremely low. This meant that more than two-thirds of the DNA in the bone had come from the dead individual! By comparison, only 4 percent of the DNA from our very best Neanderthal remains did so; more typically, the proportion was well below 1 percent. This bone was as well-preserved as the mammoth that Hendrik Poinar had sequenced and the Eskimo that Eske Willerslev in Copenhagen had sequenced. But both of those specimens had been deep frozen in the permafrost shortly after death. This explained why the majority of the DNA in those specimens was not bacterial, but I could not explain why the individual from Denisova Cave had produced so much DN
A. Whatever the reason, it certainly made the analysis of the genome much easier. In fact, our biggest issue was how to weed out the microbial DNA fragments in the library rather than how to fish out the few endogenous DNA fragments, as we had done with the Neanderthals. Now the major question was a good one: Just how much of the nuclear genome could we get? As always, we didn’t want to use the outermost surface of the bone fragment. First, it seemed irresponsible to use it all up, as we didn’t know how much of the larger piece Eddy and his group had used up in Berkeley. Second, if any part of the bone was contaminated from people handling it, it would be the surface. So Johannes used the internal part of the bone to produce two extracts. From test runs of libraries prepared from these DNA extracts, Martin Kircher calculated that we would be able to get even more coverage of the genome than we had for the Neanderthals.

  When Johannes made libraries from the extracts, he applied one of Adrian Briggs’s innovations to deal with the chemical damage that changed C nucleotides in the DNA to U nucleotides. Adrian had shown that most of these U nucleotides were found close to the ends of the ancient DNA molecules, and how to remove the damaged ends. In doing so, he lost an average of one or two nucleotides at the ends of about half the ancient molecules but he also got rid of the vast majority of errors in the DNA sequences. Since it was no longer necessary to take frequent C to T errors into account, the mapping of the fragments to the human genome became easier. Johannes made two large libraries with this method. Not only were about 70 percent of the DNA fragments in those libraries from the Denisova individual, but those DNA fragments now carried many fewer errors than the Neanderthal DNA fragments. This was real progress. Yet, I was nervous, knowing that Eddy’s group might also be at work on the same project, or even polishing a nice manuscript presenting the genome. So I tried to get everything to move as fast as possible, asking the sequencing groups to set other projects aside and sequence these libraries as fast as they could.

  I was also very curious about the strange-looking tooth that Anatoly had given us. Only DNA work would tell us if it came from the same type of person as the finger bone. Johannes, as careful as any dentist treating a live patient, drilled a small hole in the tooth and made extracts from the powder he retrieved and, in turn, libraries from the DNA in the extracts. From the libraries he then fished out mtDNA fragments. In addition, we immediately sequenced random DNA fragments from the libraries to see how much of the DNA was endogenous to the individual.

  There was good news and bad news. The good news was that he was able to reconstruct the entire mtDNA genome. There were two differences between it and the finger bone, which meant both that it was from a different person and that they were the same type of humans. The bad news was that the fraction of endogenous DNA in the tooth was only 0.2 percent. We were now even more mystified about why the finger bone contained so much endogenous DNA. I speculated that the finger might have rapidly desiccated after death, which might have limited the degradation of the DNA by enzymes in the dying cells and stopped bacterial growth. I joked that perhaps this person died with her pinky pointing up into the air so that it mummified before bacteria had too much of a chance to multiply.

  Now that we had shown that the tooth came from the same type of human as the finger, Bence devoted himself to the analysis of its morphology with renewed energy. Although I am no tooth expert, even I found it to be startlingly large. It was almost 50 percent larger than my molars. Bence pointed out that besides being very big, it was different from most Neanderthal molars with respect to both the absence and presence of certain traits in its crown. Also, its roots were unusual. Unlike Neanderthal molar roots, which tend to be closely spaced or even fused, it had strongly diverging roots. Bence concluded that the tooth morphology suggested that the Denisova population was distinct from both Neanderthals and modern humans. In fact, since the Denisova tooth lacked Neanderthal features that evolved about 300,000 years ago, he surmised that the ancestors of Denisova individuals had gone their separate way from Neanderthals before that. This was in line with what the mtDNA told us. But I was always cautious, some might even say overly skeptical, about the interpretation of morphological traits. Perhaps the Denisova people had reverted to having ancient-looking teeth after separating from either modern humans or Neanderthals. Only the nuclear genome would tell the complete story.

  Our sequencing machines began churning out Denisova nuclear DNA sequences at around the same time that we were dealing with the reviewers’ comments and finalizing the Neanderthal paper. Thus, we didn’t have much time to look at the Denisova sequences immediately, but I imagined that we could analyze them quickly once we got to it. During the last four years we had developed computer programs to analyze the Neanderthal genome that could now be directly applied to the genome from the Denisova individual. Still, I remained afraid that Eddy might be far ahead of us, so I decided to scale down the Neanderthal Genome Analysis Consortium to a core and, I hoped, faster group, asking them to devote their full attention to the Denisova genome. Most crucially, we needed David Reich, Nick Patterson, and Monty Slatkin and his crew (see Figure 23.1). We initially called ourselves the “X-Man” group because we didn’t know what the Denisova individual was. Bence had by then told us that the finger was from a young individual, perhaps just three to five years old, and we had sequenced the maternally inherited mtDNA so it seemed inappropriate to use a designation that made everyone think of a macho comic figure. I considered “X-Girl” but thought that sounded too much like a Japanese manga character. Finally, I settled on “X-Woman”—and the name stuck. Right away, the X-Woman Consortium began having weekly phone meetings.

  Udo mapped the DNA fragments to the human and chimpanzee genomes. It was comparatively easy given that we had used Adrian’s approach to remove the majority of the errors, but Udo warned me that the mapping was preliminary. In spite of this, we distributed the data to the X-Woman Consortium. Not long after we had submitted the final version of the revised mtDNA paper to Nature, Nick Patterson sent me a report on its preliminary analysis of Udo’s preliminary mappings. When I read it, I felt grateful to the reviewer who had convinced us not to name a new species. Nick had found two things.

  First, he found that the nuclear genome of the Denisova finger bone was more closely related to the Neanderthal genome than to the genomes of people living today. In fact, it seemed to be only slightly more different from the Neanderthal genome than the deepest differences one could find among humans living today—for example, between the Papua New Guinean individual we had sequenced and the African San individual. This was quite a different picture than the one painted by the mtDNA results alone, and my immediate suspicion was that gene flow from some other more ancient hominin in Asia was responsible for introducing the mtDNA into the Denisova individuals. After all, we had just shown that modern humans had interbred with Neanderthals, so gene flow seemed a reasonable guess. But it was something we needed to think carefully about.

  Figure 23.1. Monty Slatkin, Anatoly Derevianko, and David Reich, at a meeting at Denisova Cave in 2011. Photo: B. Viola, MPI-EVA.

  The second thing Nick had found was even more unexpected. Among the five humans we had sequenced for the Neanderthal analysis, the Denisova individual shared more derived SNP alleles with the Papuan individual than with the Chinese, European, or two African individuals. One possible explanation was that relatives of the Denisova individual had mixed with the ancestors of the Papuan individual, although given the distance from Siberia to Papua New Guinea I felt we might be jumping to conclusions. There could be some systematic error in what we did, and Udo again warned me that his mappings of the DNA fragments to the genome were preliminary. Perhaps there was something in the complex computer analyses that created extra similarity both between the Denisova and Neanderthal genomes and between the Denisova and Papuan genomes. Then both of Nick’s findings could be wrong.

  A week later Ed finished his own careful analysis of the new data. He found that there were very few
Y chromosomal fragments among the DNA we had sequenced, so X-Woman really was a woman, or rather, given the tiny bone, a girl. The general lack of Y chromosomal fragments also indicated that male nuclear DNA contamination was low. When he looked at divergence of the Denisova DNA sequences from the human and Neanderthal genomes, he, like Nick, found that the Denisova genome shared more derived SNP alleles with the Neanderthal genome than with modern humans. So this suggested that the common ancestor of the Denisova girl and Neanderthals first diverged from the lineage that includes modern humans, and only then did the ancestor of the Denisova girl and Neanderthals go different ways. In other words, the Denisova girl and Neanderthals were more closely related to each other than they were to modern humans. Several questions arose as we discussed these data during our Friday meetings in Leipzig and during long phone meetings with Nick, David, Monty, and the others. How could the Denisova mtDNA be so different when the Denisova nuclear genome was closer to Neanderthals than to modern humans? Could the Denisova girl perhaps have had recent ancestors who included Neanderthals and some more archaic human form, perhaps late Homo erectus? Or could she be a mixture of modern humans and such an archaic hominin? We looked at each of these possibilities and none seemed to fit.