Neanderthal Man (13 page)

I worried for months over how to go about this. I knew all too well how tricky it could be to work with museum curators, who were entrusted  with the difficult task of preserving valuable specimens for future generations while at the same time facilitating research. I had found, in some instances, that they considered their primary role to be the exertion of power, refusing access to specimens even when the possible gain of knowledge seemed to far outweigh the value of preserving a small piece of a bone. If such curators were approached in the wrong way, they could say no and then, for all-too-familiar reasons of human pride, find it difficult to go back on their word. While fretting about this, I received one day, in the most remarkable of convergences, a phone call from Bonn. The caller was Ralf Schmitz, a young archaeologist who, together with the Bonn museum curator, was responsible for the Neanderthal type specimen. He asked if I remembered an exchange we had had a few years back.

He reminded me that in 1992 he had asked what the chances of success were if one were to try to obtain DNA from a Neanderthal. This conversation had slipped out of my mind, being one out of many with archaeologists and museum curators. Now I remembered. At the time I had not known what to answer. My immediate, and slightly delinquent, impulse had been to suggest that the chances were good, so that they might readily part with a Neanderthal bone. But almost as quickly I had realized that honesty was the best way forward. After some hesitation, I had said that in my opinion we might have a 5 percent chance of success. Ralf had thanked me and I had heard nothing more from him since.

Now, almost four years later, Ralf was on the phone and said that, yes, we would be allowed to have a piece of the Neanderthal from Neander Valley. As it turned out (Ralf later told me), others had approached the museum to request samples, saying they were almost certain to get usable DNA out of the specimen. The museum authorities had then prudently decided to get the opinion of another lab and had asked Ralf to contact me. Not only our track record but also our apparent honesty in suggesting that the chances were slim had convinced Ralf and the museum that we would be their best partners. They were, as it turned out, the exact opposite of the obstructionist museum curators I had been fretting about. I was delighted.

What followed were weeks of discussions with the museum about how much bone material we would get, and from which part of the skeleton. In total, there was about half of a skeleton of what seemed to be a male individual. Our experience had shown us that the best chances of success were with compact bone—for example, a part of the shaft of an arm or leg bone, or the root of a tooth, rather than thin bones with a large marrow cavity, such as ribs. Eventually we agreed on a piece of the right upper arm,  from a part where the shaft had no ridges or other features of interest to paleontologists, who study how muscles had attached to the bone. It also became clear that we would not be allowed to remove the sample ourselves. Ralf and a colleague came to see us in Munich, and we gave them a sterile saw, protective clothing, sterile gloves, and containers in which to store the sample—and off they went. In the end, it was probably fortunate that I was not allowed to put the saw to the archetypal Neanderthal myself. I would probably have been too intimidated by this iconic fossil and would have cut off a very small piece, perhaps too small for success. When we received the sample, we were impressed by the size of what they had removed—3.5 grams of what looked like very well-preserved whitish bone (see Figure 5.3). Ralf reported that when they sawed through the bone, a distinct smell of burnt bone spread through the room. This, we believed, was a good sign; it had to mean that collagen, the protein that makes up the matrix of bone, had been preserved. It was with awe and trepidation that I approached my graduate student Matthias Krings, who had spent more than a year on fruitless attempts to extract DNA from Egyptian mummies—with the plastic bags containing the piece of the Neanderthal type specimen and asked him to apply our latest and best methods to it.

Figure 5.3. The right upper arm bone of the Neanderthal type specimen with the sample removed by Ralf Schmitz in 1996. Photo: R. W. Schmitz LVR-LandesMuseum Bonn.

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During the weeks and months after our publication of the Neanderthal mtDNA sequence, I reflected on what had led up to it. I had come a long way from my first attempts sixteen years earlier to extract DNA from a piece of dried calf’s liver from the supermarket. Now, for the first time, we had used ancient DNA to say something new and profound about human history. We had shown that the archetypical Neanderthal carried mitochondrial DNA very different from the mtDNA in people today, and that he or his relatives had not, before they became extinct, contributed their mtDNA to modern people. The achievement had required years of painstaking work to develop techniques to reliably determine DNA sequences from individuals long dead. Now that I had these techniques at my disposal, and a group of dedicated people able and willing to try new things, the biggest question was: Where should we go from here?

One task seemed of immediate importance: to determine mitochondrial DNA sequences from other Neanderthals. As long as we had studied only one individual, it remained possible that other Neanderthals carried mitochondrial genomes very different from the one from Neander Valley, perhaps even carrying mitochondrial genomes that were like those of present-day humans. Mitochondrial DNA sequences from additional Neanderthals would also reveal something about the genetic history of the Neanderthals themselves. Present-day humans, for example, have relatively little genetic mtDNA variation. If Neanderthals did, too, this would suggest that they had originated and expanded from a small population. If, on the other hand, they had as much mtDNA variation as any of the great apes have, this would suggest that over their history their numbers had never been very low. They would not have had such a dramatic history with ups and downs in population size as modern humans have. Matthias Krings, eager to follow up on his success with the iconic type specimen from Neander Valley, was keen to examine other Neanderthal specimens.

The major problem was getting access to fossils sufficiently well preserved for us to do work.

I thought a great deal about why we had been successful with the Neander Valley type specimen and came to realize that the fact that it had come from a limestone cave might be significant. Tomas Lindahl had taught me that acid conditions cause DNA strands to disintegrate, which was why the Bronze Age people found in acid bogs in northern Europe had never yielded any DNA. But when water passes over limestone, it becomes slightly alkaline. So I decided we should concentrate on Neanderthal remains unearthed in limestone caves.

Unfortunately, I had never paid much attention to the geological features of Europe in school. But I remembered the first anthropological conference I had ever attended, in Zagreb, in what was then Yugoslavia, in 1986. During the conference, we were taken on excursions to Krapina and Vindija, two sites where large amounts of Neanderthal bones had been found in caves. I made a quick search in the literature and confirmed that both Krapina and Vindija were limestone caves, which was promising. Promising as well was the presence of large numbers of animal bones, particularly of cave bears, in the caves. Cave bears, which were a large plant-eating species, became extinct shortly after 30,000 years ago, just like the Neanderthals. Their bones often abound in caves, often in circumstances suggesting that they died during hibernation. I was happy about the presence of cave-bear bones because they could possibly serve as a convenient tool to check whether DNA was preserved in the caves. If we could show that their bones contained DNA, this might be a good means to convince hesitant curators that they should allow us to try the much more valuable Neanderthal remains from the same cave. I decided to interest myself in cave-bear history, especially in the Balkans.

After a bloody war with Serbia, Zagreb had become the capital of the independent Republic of Croatia. The largest collection of Neanderthals there is from Krapina, in northern Croatia, where starting in 1899 the paleontologist Dragutin Gorjanovi
ć
-Kramberger discovered more than eight hundred bones from some seventy-five Neanderthals—the richest cache of Neanderthals ever found. These bones are today housed in the Museum of Natural History in the medieval center of Zagreb. The other site, Vindija Cave (see Figure 6.1) in northwestern Croatia, was excavated by another Croatian paleontologist, Mirko Malez, in the late 1970s and early 1980s. He  found bone fragments of several Neanderthals but no spectacular crania like those found in Krapina. Malez also found enormous amounts of cave-bear bones. His finds are housed in Zagreb, too, in the Institute for Quaternary Paleontology and Geology, which belongs to the Croatian Academy of Sciences and Arts. I arranged to visit both this institute and the Museum of Natural History. In August 1999, I arrived in Zagreb.

The Krapina Neanderthal collection was extremely impressive, but I was skeptical about its potential for DNA research. The bones were at least 120,000 years old and therefore older than anything we had found to yield DNA. The Vindija collection looked more promising. First of all, it was younger. Several layers in the excavation had yielded Neanderthal remains, but the uppermost and thus the youngest one to do so was between 30,000 or 40,000 years old—young, as far as Neanderthals go. I saw a second exciting feature of the Vindija collection: it was full to overflowing with ancient cave-bear bones. They were stored, according to bone type and layer, in innumerable paper sacks that were coming apart in the humidity of the Quaternary Institute’s basement. There were sacks full of ribs, others full of vertebrae, others of long bones, and yet others of foot bones. It was an ancient DNA gold mine.

Figure 6.1. Vindija Cave in Croatia. Photo: J. Krause, MPI-EVA.

In charge of the Vindija collection was Maja Paunovic, a woman of a certain age who spent her days in an institute without public exhibitions and with few facilities for doing research. She was friendly enough but understandably dour—no doubt aware that science had passed her by. I spent three days with Maja, going through the bones. She gave me cave-bear bones from several layers at the Vindija site as well as small samples of fifteen of the Neanderthal bones. This seemed exactly what we needed for the next step in our exploration of the genetic variation among Neanderthals. When I flew back to Munich I felt confident that we would make quick progress.

In the meantime, Matthias Krings had extended his sequencing of the Neanderthal type specimen to a second part of the mitochondrial genome. The results confirmed that this specimen’s mitochondrial DNA shared a common ancestor with present-day human mtDNAs about half a million years ago. But this was of course what we had expected, so the news felt slightly boring after the emotional high produced by the first Neanderthal sequences. Not surprisingly, he was eager to throw himself on the fifteen Neanderthal bone samples I had gotten from Maja in Zagreb.

We first analyzed their state of amino-acid preservation. Amino acids are the building blocks of proteins and can be analyzed from much smaller samples than are needed for DNA extractions. We had shown before that if we could not find an amino-acid profile suggesting that the samples contained collagen (the main protein in bones), and if the amino acids were not present largely in the chemical form in which they are built into proteins by living cells, then our chances of finding DNA were very small and there would be no point in destroying a larger piece of the bone in an attempt to extract it. Seven of the fifteen bones looked promising, with one that particularly stood out. We sent a piece of that bone for carbon dating and the result showed that it was 42,000 years old. Matthias made five DNA extracts and amplified the two mitochondrial segments he had studied in the type specimen. It worked nicely. He sequenced hundreds of clones, taking pains to ensure that every position was observed in at least two amplifications that, at my insistence, should come from different extracts, in order to make absolutely sure that they were totally independent of each other.