Cave bear genomics
A new article on the epub area of Science, by James Noonan (Lawrence Berkeley National Laboratory) describes the recovery of nuclear DNA sequences from cave bear remains. Here's the abstract:
Despite the greater information content of genomic DNA, ancient DNA studies have largely been limited to amplification of mitochondrial sequences. We describe metagenomic libraries constructed using unamplified DNA extracted from skeletal remains of two 40,000-year-old extinct cave bears. Analysis of ~1 Mb of sequence from each library showed that, despite significant microbial contamination, 5.8% and 1.1% of clones contain cave bear inserts, yielding 26,861 base pairs of cave bear genome sequence. Comparison of cave bear and modern bear sequences revealed the evolutionary relationship of these lineages. The metagenomic approach employed here establishes the feasibility of ancient DNA genome sequencing programs.
In a previous post, I covered the recent announcement that this group -- a collaboration between Max-Planck Evolutionary Anthropology and Lawrence Berkeley lab -- plans to recover genomic sequences from Neandertals. The cave bear paper gives a clear hint about how it will be done.
A Nature news article covers the bear research:
The standard practice for sequencing genes involves making numerous copies of the initial sample through a process called a polymerase chain reaction, or PCR. Subjecting ancient DNA to this does not produce good results because PCR picks up and duplicates the sequences of modern animals more efficiently. This means that bits of contaminating DNA often drown out samples from the prehistoric animal.
"The prevailing idea was that this was impossible," says James Noonan of the Lawrence Berkeley National Laboratory in California, who is lead author of the paper that appears in Science this week.
To overcome this challenge, Noonan and his colleagues decided to skip the replicating step and directly sequence the tiny amount of DNA extracted from two Austrian cave-bear bones that are more than 40,000 years old. To make sure each portion of DNA was really from the bears rather than a contaminating source, they compared each sequence produced with the genome of the dog, a modern relative of the bear.
The technologies needed to examine such tiny amounts of DNA directly, along with the reference genome from the dog, have become available to scientists only recently.
The team determined that nearly 6% of the sequences analysed from one of their animal samples belonged to ancient bear: an unexpectedly large amount. The rest of the DNA probably came from soil microbes or the palaeontologists handling the bones, the team says.
Metagenomics
The technique they are using, called metagenomics, is borrowed from environmental science. The principle is that you take a sample of organic material and look for evidence of the organisms within it by separating out all the DNA and cloning it.
This is in contrast to PCR, where you look for a specific piece of DNA from one location in the genome by designing primers that will amplify that piece preferentially. With metagenomics, you don't start out knowing what you are looking for.
Metagenomics is useful to environmental scientists, drug researchers, and others because it allows the study of DNA from organisms without being able to culture the organisms in the laboratory. You are taking DNA from the samples and inserting it into bacterial colonies using a vector, resulting in a "metagenomic library." This library consists of DNA fragments from any kind of organisms that were in the sample, possibly including hundreds of species. If you've heard of the idea of creating a "bar code" of DNA that could identify organisms taken from ocean water or soil samples, this is the science that is behind that idea. You don't know what you're extracting from, and you'd like a way to standardize samples so you can say.
For the cave bears, what has been done is the extraction of DNA from the sample and cloning into a metagenomic library, consisting of bacterial DNA, fungal DNA, human DNA, and some cave bear DNA. Then the lab sequences the cloned fragments to find out what they are. The ones that look bear-like, they assume are endogenous. Hence, a limitless source of cave bear genetic material.
Of course, in the case of the bears, the lab has little worry that living bears in the laboratory have handled and contaminated the remains (although I have seen cases in labs where such strange contaminations have happened...). For Neandertals, the possibility of human contamination is everpresent. That this technique skips the PCR step is very important in limiting contamination (since modern DNA amplifies much more readily than ancient DNA) but it far from eliminates the problem. The two cave bear extracts preserve a substantial amount of human sequence -- in one case a third as much human contaminant as original cave bear. It will be very hard to exclude this contamination from consideration in a Neandertal extract, which is very likely to share much of its genome in common with humans without contamination.
Why did they compare with dogs? Because there is a dog genome project, but not a bear one. This is a computational comparison, not a wet one. For Neandertals, the comparison will be the same: hunting through the human genome to find segments that correspond to the Neandertal extracts.
Looking for Neandertal genomic DNA
This is new stuff, to a point, but not all that new. The original extraction of Neandertal mtDNA in 1997 used bacterial cloning to reconstruct the fragments. The history indicates that Pääbo's lab has not trusted PCR amplification in Neandertal-aged remains from the beginning, and certainly for good reason considering the very high chance of preferential amplification of contaminants.
But the metagenomics approach adds a new twist. If you aren't looking specifically for one genomic region when you extract DNA from the sample and clone it, then the results are going to be a scatter from across the genome. In this case, Neandertal genomics may really be like Forrest Gump's box of chocolates: you never know what you're going to get. With a sufficiently large sample, you could in principle find any region of the genome. But it's not obvious how much extract a sufficiently large sample would take. For the bears, around 1 megabase was cloned, yielding around 27 kilobases of cave bear DNA. With more effort, a larger quantity might be obtained, but of course this would require the destruction of larger samples of bone.
Twenty-seven kilobases is a potentially interesting amount. It is large enough to give a good chance of finding genetic variants in the Neandertal sequence. Humans vary in around 1 nucleotide for every thousand, so 27 kb is a nice chunk of potential differences.
But if only one out of a thousand base pairs are different between humans, the amount of DNA degradation over time might overwhelm the actual number of changes. There is some evidence from ancient mtDNA sequences for diagenetic damage to the preserved sequences resulting in sequence changes. These are known to be diagenetic because some of them apparently occur at predictable hotspots, but the rate of this damage is not yet known, and it appears to differ between different specimens. Nuclear DNA may be more stable than mitochondrial DNA, because it is packaged by proteins into a firmer structure, but I wouldn't make any bets on it. But even so, this process of diagenetic change has the potential to be much greater than the actual rate of evolutionary differences. So it will be a terrible problem to interpret the genetic differences.
Noonan et al. (2005:3) observe this problem in the cave bear sequence:
The substitution rate we estimated for cave bear is higher than that in any other bear lineage. On the basis of results from PCR-amplified ancient mitochondrial DNAs, cytosines in ancient DNA can undergo deamination to uracil, which results in an excess of G to A and C to T (GC-AT) transitions (22). The inflated substitution rate in cave bear is likely due to an excess of such events, since many of the substitutions assigned to the cave bear lineage are GC-AT transitions (Fig. 3A). These presumably damage-induced substitutions complicate phylogenetic reconstruction and the identification of functional sequence differences between extinct and modern species.
They argue that the diagenetic changes may be excluded if they occur in a subset of the clones, as they apparently do in this case. They merely leave out the clones with high rates of GC-AT transitions, and their results look more normal. This helps to reduce the problem, if the changes are concentrated in certain clones, but it cannot eliminate it.
This might be easier if we knew we were looking for particular variants at certain genomic locations. For example, if the lab went looking for the FoxP2 gene, they could expect to find variation at the one or two amino acid changing substitutions that have occurred in humans compared to chimpanzees. The odds of diagenetic changes at these positions would be relatively low compared to the known odds of finding a genetic substitution there. But the metagenomic approach may not give the opportunity to focus in on changes that are known to be likely polymorphisms. We may have to just take what we can get.
In any event, it should be interesting to see these results come out. I am afraid that we will see phylograms showing the relationship of some Neandertals compared to other living human populations. That would be a mistake, since living people are not related as branches on a tree; and there is no necessary reason to suppose that Neandertals were either. But I guess that's my job to point out when the time comes.
References:
Noonan JP, et al. 2005. Genomic sequencing of Pleistocene cave bears. Science Express. doi: 10.1126/science.1113485. Abstract
FOXP2 is really recent, it really did introgress (if it's not contamination)
That's the thrust of a technical comment by Graham Coop and colleagues, now online in Molecular Biology and Evolution. The letter refers to the extraction of FOXP2 from two Neandertal specimens from El Sidrón, by Johannes Krause and colleagues, reported last year (I wrote about the paper here).
First, the bad news. The current letter raises the prospect of contamination. Notably, the controls applied by Krause et al. (2007) may be relatively weak evidence against contamination, because of polymorphism within large human comparative samples. The tests rely on the assumption that there is little DNA from living humans in the samples. But if we cannot distinguish Neandertal from human DNA with great accuracy, then we will be mistaken some proportion of the time. Krause et al.'s test, based on derived human alleles absent from the Neandertal genome draft, can still go wrong if the human contaminants happen to have all the ancestral (non-derived) human alleles.
Well, that seems to be the story these days with Neandertal DNA extraction. No test of contamination is good enough. (And remember, that every "test" of contamination is really a procedure for excluding the hypothesis that ancient sequences are identical to recent ones.)
Now, the more interesting news. Coop and colleagues verify that the selective sweep affecting human FOXP2 was indeed recent -- they estimate 42,000 years ago:
To demonstrate this, we estimated the time of the most recent common ancestor (tMRCA) of the selected haplotype (see Figure 1), using an approach sometimes called phylogenetic dating (Thomson et al. 2000; Hudson 2007). This method does not make assumptions about demography and selection, but only requires that the mutations in the intron be neutral or nearly neutral. Taking this approach, we obtained a mean tMRCA of 42 Kya (see SOM for details). While there is considerable uncertainty associated with this estimate, it is surprisingly recent if selection took place over 300 Kya (see SOM). In other words, the selective scenario proposed by the authors cannot account readily for patterns of variation in modern humans. Given that we have no power to detect a beneficial substitution that occurred over 250 Kya, (cf. Sabeti et al. 2006) yet we see a footprint of positive selection at FOXP2, the conclusion of a recent selective sweep at FOXP2 is not surprising (Coop et al. 2008:3-4).
FOXP2 is in one of the ENCODE regions, so its variation is pretty well known. This is not a problematic case: it has a very limited amount of variation around it, and has a strong excess of rare alleles, both signs of a recent sweep.
Coop and colleagues suggest that the beneficial human allele spread into Neandertals (or vice versa) by low levels of gene flow coupled with its selective advantage -- in other words, introgression.
They do allow for an alternative -- perhaps the two amino-acid-coding mutations were not the target of selection, but instead some linked locus. This would not erase the necessity of gene flow from Neandertals, but would question whether this gene flow had involved the FOXP2-language scenario, since it might be some linked gene unrelated to language.
(CORRECTION (2008/04/18): If selection were on a linked site, then Neandertals might share the human-derived amino acids as a result of ancient shared ancestry with humans, while the linked selected sweep might be absent in Neandertals, not necessitating any gene flow.)
I doubt this hypothesis of a linked sweep, since the two sites with human-derived substitutions are otherwise very strongly conserved among mammals. This looks like a credible target for recent selection. But the hypothesis of selection on a linked site cannot presently be tested.
So that's the story. It seems very likely that Neandertals got the language gene from us, or us from them, long after many other genes in the two populations diverged. I write "many" rather than "most" because we haven't really been able to assess the proportion of derived alleles shared by humans and Neandertals. The completion of the draft sequence may help, but I'm afraid that the specter of contamination is going to keep on being raised whenever a part of the Neandertal draft genome looks humanlike.
(via Dienekes)
References:
Coop G, Bullaughey K, Luca F, Przeworski M. 2008. The timing of selection at the human FOXP2 gene. Mol Biol Evol (in press) doi:10.1093/molbev/msn091
Neandertal introgression, genetic-style
The paper by Patrick Evans and colleagues, from Bruce Lahn's lab, is now live (and free) at PNAS. There is a short news report by Michael Balter at ScienceNOW, and the Howard Hughes press release is admirably clear.
If you've been hearing a lot about the word "introgression" lately, this is why. At least, the first of the reasons why.
Here's the abstract:
At the center of the debate on the emergence of modern humans and their spread throughout the globe is the question of whether archaic Homo lineages contributed to the modern human gene pool, and more importantly, whether such contributions impacted the evolutionary adaptation of our species. A major obstacle to answering this question is that low levels of admixture with archaic lineages are not expected to leave extensive traces in the modern human gene pool because of genetic drift. Loci that have undergone strong positive selection, however, offer a unique opportunity to identify low-level admixture with archaic lineages, provided that the introgressed archaic allele has risen to high frequency under positive selection. The gene microcephalin (MCPH1) regulates brain size during development and has experienced positive selection in the lineage leading to Homo sapiens. Within modern humans, a group of closely related haplotypes at this locus, known as haplogroup D, rose from a single copy 37,000 years ago and swept to exceptionally high frequency (ca. 70% worldwide today) because of positive selection. Here, we examine the origin of haplogroup D. By using the interhaplogroup divergence test, we show that haplogroup D likely originated from a lineage separated from modern humans 1.1 million years ago and introgressed into humans by ca. 37,000 years ago. This finding supports the possibility of admixture between modern humans and archaic Homo populations (Neanderthals being one possibility). Furthermore, it buttresses the important notion that, through such adminture, our species has benefited evolutionarily by gaining new advantageous alleles. The interhaplogroup divergence test developed here may be broadly applicable to the detection of introgression at other loci in the human genome or in genomes of other species.
I'm starting a second post with Q and A regarding the paper.
Here, I want to note some news:
- I have my own paper on introgression (with Greg Cochran) that will be coming in PaleoAnthropology, so it is a topic that to which we've devoted a lot of consideration.
- There will be more Neandertal news in the next week. These are busy Neandertal times!
- Because we've been working on this topic, I've been avoiding it. But now that some of this stuff has come out, I will point out that there is now a substantial literature on genetic introgression from archaic humans. More in the Q and A.
UPDATE (11/8/2006): My colleague, Greg Cochran, has a post at GNXP discussing introgression and microcephalin further:
If this pans out the way we think it will, introgression from Neanderthals (and maybe with other archaics) may have been one of the two fundamental patterns underlying recent human evolution.
One of the two.
References:
Evans PD, Mekel-Bobrov N, Vallender EJ, Hudson RR, Lahn BT. 2006. Evidence that the adaptive allele of the brain size gene microcephalin introgressed into Homo sapiens from an archaic Homo lineage. Proc Nat Acad Sci (early edition) DOI link
The amazing talking Neandertals
This week, Johannes Krause and colleagues from the Max Planck Evolutionary Anthropology institute announced that they had tickled FoxP2 out of two Neandertal specimens from El Sidrón, Spain. The bones were excavated in sterile (clean-cave?) conditions, immediately frozen and then shipped to Leipzig, where extracts were taken in clean-room conditions.
Here's an FAQ about what they found.
Why is this paper really important?
Isn't it obvious? It's important because it demonstrates that more than one Neandertal is suitable for nuclear genome recovery. We will know about genetic variation in Neandertals, sooner rather than later. These two bones come from different individuals, because the Leipzig group found two different mtDNA sequences in them. Together with the Vindija Vi 33.16 specimen in the original Neandertal genome papers, this makes three nuclear genome Neandertals. There will be more.
It also shows the possibility of probing ancient skeletons for specific genes. Here, they went in looking for Y-DNA, X-DNA and particular sites on FoxP2, and they found them. That is definitely the way to go if you want to test a biologically significant hypothesis fast -- otherwise, you just have to wait until the sequence comes up in your genome project.
However, I question the value of probing for individual genetic variants in this way. Every probe takes a bit of sample, which might be more efficiently used in whole-genome sequencing. We have 25,000 genes, and every one is potentially interesting. Every small sample used to assay only one of those genes may destroy many sequences from the others. It would be one thing if samples were trivial and easily replaced, but they obviously aren't.
Still, we will certainly see additional probes for genes that are of particular interest. I wouldn't be surprised to see MC1R results soon, to probe whether there were pigmentation variants in Neandertals. The same has already been done for woolly mammoths.
So, Neandertals had the human-specific FoxP2 form. Did they talk?
I think the genetic observation leans toward that direction, but doesn't really change our understanding. Consider:
Neandertals have a hyoid bone with humanlike anatomy, as did the Atapuerca people at more than 300,000 years ago, even though A. afarensis did not. So something related to vocalization evolved in humans by the Middle Pleistocene. Although Neandertal vocal tracts may not have been identical to recent humans, there is nothing about them that would preclude speech. The only paleoneurological observation about language puts a developed Broca's area on the KNM-ER 1470 endocast, Homo habilis.
Like other Middle Paleolithic/MSA people, their technology required more information to learn than earlier, Lower Paleolithic industries, leading to regional differentiation and more task-specific facies. Late Neandertals made use of some technology otherwise used only by Upper Paleolithic modern humans. Their hunting methods must have required cooperation and may have been impossible without a more sophisticated communication strategy than used by other primates.
All of these things argue for some kind of Neandertal language irrespective of FoxP2.Then again, most of the arguments against humanlike language facility in Neandertals also have nothing to do with FoxP2, either. The slow technological progress, limited collection strategies, the rarity of any artistic or symbolic expression, their high mortality rate, and -- of course -- the fact that they no longer exist have all been considered as evidence that Neandertals lacked some essential aspect of "behavioral modernity." If language is a prerequisite for the modern human pattern of behavior, then Neandertals may not have talked, at least not in the way we do.
I think the FoxP2 story has really confused people much more than necessary. But in this case, the confusion is the same that results from every other gene study: when the press says we've found a gene "for" something, what it ought to say is that we've found an allele that affects something.
No macromutation happened. Language did not spring full-formed into the mind of some ancient African. All members of Homo used communication systems including some (possibly minimal) elements of language, and the evolution of the human brain, along with technological changes throughout the Paleolithic, reflect the evolution of communication. Human language evolved -- like all things -- over a long time, and like all complex phenotypes it required a series of mutational changes. Many of these mutations became fixed during recent human evolution, some may still be changing in frequency today. Language evolution is probably a continuing process.
That means that it must have involved many more genes than FoxP2 -- which after all experienced only two amino acid substitutions in all of human evolution. I would imagine the number of genes involved in language evolution is more than 500, and I wouldn't be surprised if it were much more. In that context, it seems quite silly to say FoxP2 is the "critical" evolutionary change for anything.
Then you agree with Language Log. They told me that FoxP2 isn't a "language gene."
The case is strong that the two FoxP2 coding substitutions in humans were selected because of their role in language. The gene sequence is strongly conserved in most mammals, and shows similar changes in some other species with unusual vocal adaptations, such as echolocating bats (Li et al. 2007). Its expression pattern delineates areas related to vocalizations in both humans and birds, and the pattern itself differentiates between song-learning versus nonlearning bird species (Haesler et al. 2004, Teramitsu et al. 2004, Webb and Zhang 2005). And of course, mutations to FoxP2 can result in specific language impairment (SLI) in humans.
Still, that case is only circumstantial. We know that FoxP2 was under selection, that it became fixed in humans, probably during the Late Pleistocene, and that breaking the gene changes brain development and damages language skills. But we don't know what a human would be like with the chimpanzee form of the protein. We don't know whether both of the human-specific amino acid substitutions have a different effect than one. Most important, we don't know what other genetic changes may have been necessary backgrounds for selection on FoxP2.
This means Neandertals were really modern humans, right?
This study should put an end to the "sudden mutation" model of modern human origins.
There was not a single mutation that made the critical difference in the ancestry of today's people. There was no cognitive Rubicon leading to modern human evolution. I would analogize the process as a slow-motion avalanche: at first a few small sands began to tumble, and then selection on a large number of genes became inevitable. FoxP2 is one of those genes, and as yet we don't know whether it was near the beginning or near the end of the process.
But it is clear that the process began before the Neandertals were gone. Some aspects of behavioral complexity did begin to evolve rapidly sometime after 70,000 years ago. This rapid evolution was multiregional in context -- it was not limited to a single human population. In particular, it was not limited to Africans: the last Neandertals clearly manifested technological and behavioral strategies otherwise defined as "behaviorally modern" (d'Errico 2003). There's a reason why the Neandertal-produced Châtelperronian industry of France and Spain was historically considered the first Upper Paleolithic industry.
But we have undergone light-years of change since the last Neandertals lived. This is not a question of "modern human origins" anymore. We can now show that living people are much more different from early modern humans than any differences between Neandertals and other contemporary peoples. I think that "modern humans" is on its way to obsolescence. What matters is the pattern of change across all populations. Possibly that pattern was initiated by changes in one region but the subsequent changes were so vast that the beginning point hardly matters.
We all know that the Neandertal genome is riddled with contamination from modern humans. Isn't the null hypothesis that we have a modern human sequence here because it is a modern human?
Well, as you know, I'm not all that convinced that contamination explains the interpretive discrepancies between last year's genome papers. But still, this study has done some things to address the problem of contamination.
It is notable that Green et al. (2006) found 25% modern human mtDNA in one of the El Sidrón bones: this shows that even "sterile" excavation, immediate freezing and extraction under clean-room conditions cannot exclude contamination. There is at the moment nothing more that can be done. We will always have the problem of a contamination fraction in ancient Neandertal skeletons. So we have to judge each study by the extent to which we can exclude contaminants with statistical analysis.
For this study, Krause et al. (2007) developed a test of nuclear DNA contamination: they identified seven gene variants that differ between the recovered Vindija Vi 33.16 nuclear genome and all known living humans. In other words, these are human-derived mutations that are absent from the only known Neandertal nuclear genome. Then, they probed the El Sidrón bones for these sites. They found only the ancestral form in their extracts of both bones -- presumably because no human contaminants were present in their samples.
That seems like a pretty good indication that the other sites in their sample represent the true gene variants of the ancient Neandertals. I wouldn't go so far as to say that contamination is ruled out, but it seems like these are good results.
Did FoxP2 introgress into Neandertals?
It sure looks that way to me. Let's consider the evidence:
FoxP2 recently fixed in humans. According to Enard et al. (2002:871):
Under a model of a randomly mating population of constant size, the most likely date since the fixation of the beneficial allele is 0, with approximate 95% confidence intervals of 0 and 120,000 years.
Now, Enard et al. (2002) noted that human populations have grown over time, and that they are not randomly mating, so that this date estimate might be too recent. Allowing for population growth since "10,000--100,000 years ago," they asserted that fixation of FoxP2 must have happened "during the last 200,000 years of human history." But this is not quite accurate. Unlike genetic drift, positive selection can and often does fix genes rapidly in a growing population. It simply doesn't matter that the human population has been rapidly growing: FoxP2 may still have just become fixed yesterday.
Last year, Green and colleagues (2006) considered that the Neandertal-modern population divergence time might have been quite recent, depending on the ancestral population size. According to the estimates of Wall and Kim (2007), the Green et al. data are consistent with a Neandertal-modern population divergence time as recent as 30,000 years ago. Of course, that date would predict substantial admixture between contemporary Neandertal and non-European populations -- they would have been exchanging genes up to the very lifetimes of the last Neandertals. According to those data there would be nothing surprising about Neandertals and living people sharing the human-derived FoxP2 allele. But as mentioned above, Wall and Kim (2007) used the recent divergence estimate as evidence that the Neandertal genome data from Green et al. must be contaminated.
So, if we cannot trust the data, then we have to fall back on some other estimate of the divergence date. Noonan and colleagues (2006) estimated a divergence date between Neandertals and modern populations between 170,000 and 570,000 years ago. If we accept that, then the confidence intervals of the Neandertal-human divergence and the FoxP2 selective sweep might barely overlap. Might. But I will note that a minimal overlap between the 95% confidence intervals of two point estimates does not mean that they are not significantly different. Only if the expected value of one estimate falls within the 95% confidence interval of the other do they fail to be significantly different. It is pretty unlikely that the most recent FoxP2 sweep is older than 170,000 years ago and the Neandertal-modern population split is as recent as 170,000 years.
That is, unless the "split" time reflects widespread genetic introgression.The current paper (Krause et al. 2007) goes to some contortions to try to establish that the FoxP2 sweep could really have been older than 300,000 years ago (where they place the lower confidence limit on the N-M split):
The third scenario is that the selective sweep started before the divergence of the ancestral populations of Neandertals and modern humans around 300,000-400,000 years ago
Let me just say that I was surprised to read this explanation in a paper from this group. One of the main arguments they have been posing as a scientific value of the Neandertal genome sequencing is that conventional methods don't detect selection at 300,000-400,000 years ago. But here, they consider such an ancient mutation to be the most likely hypothesis. This seems like quite a shift just to avoid the unpleasant idea of Neandertal introgression. Ooooh -- can't have those Neandercooties!
In reality, there is no reason to think the fixation of FoxP2 happened as early as 300,000 years ago, and indeed the very high frequencies of the linked derived alleles (over 97% for six of the linked alleles) suggest strongly that the sweep probably happened within the last 100,000 years -- otherwise, subsquent genetic drift should have caused these linked derived alleles to show more dispersion in their current frequencies. The same features that make the inference of selection so strong at FoxP2 -- it is far (>286 kilobases) from the nearest gene and it includes many high-frequency derived alleles in addition to reduced polymorphism -- make it very unlikely that the selective sweep was ancient.
So, considering that the El Sidrón samples both share the human-derived amino acid substitutions on the same haplotype as modern humans, complete with all the high-frequency derived SNPs, it seems almost certain that the gene introgressed into Neandertals from modern humans.
Or, there's one other option. One of the El Sidrón bones includes a relatively rare (in humans) ancestral SNP allele at one of those linked sites where the derived allele is at very high frequency in humans. One explanation: the selected mutation arose in Neandertals and introgressed into other humans. That would explain why this Neandertal didn't have a SNP variant on its FoxP2 haplotype that later became very common in humans: Neandertals had the new FoxP2 first.
What about that Y chromosome thing?
The El Sidrón bones both tested positive for the Y chromosome site assayed in the study. That means they were both male (duh!). But more important, the Y chromosomes of both individuals lacked the human-specific derived mutation that the researchers tested for. Since all human males yet surveyed have this human-derived mutation, this means that a Y chromosome variant has fixed in modern humans that Neandertals did not have. Since the entire nonrecombining portion of the Y chromosome is completely linked, we can infer that the entire modern human Y chromosome has undergone at least one fixation not shared with the ancestors of these Neandertals.
Here's the text (from page 2):
Both Neandertals yielded products for Y chromosomal primer pairs, indicating that they were males. Strikingly, all 15 Y chromosomal products for the five assayed positions show the ancestral allele. This includes two polymorphisms that define the deepest split among current human Y chromosomes (Y2 and Y4, Figure S1) as well as two polymorphisms that cover less common African Y chromosomes (Y3 and Y5, Figure S1). These Y chromosome results must derive, then, either from Y chromosomes that fall outside the variation of modern humans or from the very rare African lineages not covered by the assay (Figure S1). For our purposes, this result shows that neither the maternally inherited mtDNA nor the paternally inherited Y chromosome shows evidence of gene flow from modern humans into Neandertals or of subsequent contamination of their mortal remains.
That's not such a big surprise. Already we knew that the fixation of the human Y chromosome was very recent -- probably within the last 70,000--100,000 years, and possibly even more recently. Every man on earth shares recent Y chromosome mutations that were completely absent in Middle Pleistocene humans. That is one radical recent evolutionary change.
The paper elsewhere suggests that this absence of the human-derived Y chromosome in Neandertals as evidence that they did not contribute other genes to us. I could not disagree more.
The very recent fixation of the Y chromosome in an expanding human population is extremely unlikely to have resulted from genetic drift. Drift does not eliminate rare variants as quickly in an expanding population. Instead, one or more Y chromosome mutations must have been positively selected, resulting in the fixation of the entire NRCY in recent humans.
In that context, the Neandertal result is quite expected: they had an earlier Y chromosome lacking one or more mutations later selected in the other ancestors of living people.
References:
Briggs AW, Stenzel U, Johnson PLF, Green RE, Kelso J, Prüfer K, Meyer M, Krause J, Ronan MT, Lachmann M, Pääbo S. 2007. Patterns of damage in genomic DNA sequences from a Neandertal. Proc Nat Acad Sci USA doi:10.1073/pnas.0704665104
d'Errico F. 2003. The invisible frontier. A multiple species model for the origin of behavioral modernity. Evol Anthropol 12:188-202. doi:10.1002/evan.10113
Green RE, Krause J, Ptak SE, Briggs AW, Ronan MT, Simons JF, Du L, Egholm M, Rothberg JM, Paunovic M, Pääbo S. 2006. Analysis of one million base pairs of Neanderthal DNA. Nature 444:330-336. doi:10.1038/nature05336
Haesler S, Wada K, Nshdejan A, Morrisey EE, Lints T, Jarvis ED, Scharff C. 2004. FoxP2 expression in avian vocal learners and non-learners. J Neurosci 24:3164-3175. doi:10.1523/JNEUROSCI.4369-03.2004
Krause J, Lalueza-Fox C, Orlando L, Enard W, Green RE, Burbano HA, Hublin J-J, Bertranpetit J, Hänni C, Fortea J, de la Rasilla M, Rosas A, Pääbo S. 2007. The derived FoxP2 variant of modern humans was shared with Neandertals. Curr Biol 17:1-5. doi:10.1016/j.cub.2007.10.008
Li G, Wang J, Rossiter SJ, Jones G, Zhang S. 2007. Accelerated FoxP2 Evolution in Echolocating Bats. PLoS ONE 2(9): e900. doi:10.1371/journal.pone.0000900
Noonan JP, Coop G, Kudaravalli S, Smith D, Krause J, Alessi J, Chen F, Platt D, Pääbo S, Pritchard JK, Rubin EM. 2006. Sequencing and analysis of Neanderthal genomic DNA. Science 314:1113-1118. doi:10.1126/science.1131412
Wall JD, Kim SK. 2007. Inconsistencies in Neanderthal genomic
DNA sequences. PLoS Genet 3:e175. doi:10.1371/journal.pgen.0030175.eor
Introgression and microcephalin FAQ
Considering the paper by Evans and colleagues, I've come up with a list of questions and answers:
What is introgression?
Introgression is the transfer of alleles across species or subspecies boundaries. In other words, it describes gene flow between populations that are partially isolated. For archaic humans, there is no test of the strength or permeability of boundaries between populations; it is common to use the term "introgression" to describe gene flow in such situations, even if such gene flow is fairly common.
The paper by Evans and colleagues describes a scenario of adaptive introgression. In such cases, an allele with a selective advantage moves from one population to another.
Adaptive introgression must be a very unusual event, right? I mean, I've never heard of it before!
If you haven't heard of adaptive introgression, you haven't been reading the literature. Adaptive introgression across species boundaries is very common in mammals, and is almost ubiquitous where closely related species are sympatric. It has long been known to happen on the basis of morphological characters that spread through hybrid zones into adjacent populations. But now that molecular surveys have become common, introgressive genes have been found moving out of current hybrid zones, and also in the areas where hybrid zones likely occurred long in the past.
Hybrid zones themselves are often quite obvious. But introgression is not about hybrids. It occurs when backcrosses spread alleles into the other parental species. Hybrids may have a mixture of many genes and characters. Introgression involves a small number of genes, which are much more likely to spread if alleles are adaptive. Where different populations are in reproductive contact, adaptive introgression may often be the most important source of adaptive alleles -- it provides a way for a species or population to benefit from the adaptive evolution of neighboring species.
There is one thing that impedes introgression: linkage to deleterious alleles. Species separated for longer times are more likely to have alleles that are bad on the genetic background of related species, and so potential adaptive alleles must have advantages outweighing all the deleterious alleles they are linked to. In these situations, adaptive introgression may only occur after enough recombination has broken the adaptive allele apart from some or all of its linked deleterious neighbors.
But I thought that "species" means "no interbreeding!"
Get with the times, man! Mammal species just don't establish reproductive barriers very quickly. Comparing mammals, postzygotic isolating mechanisms take between 2 and 10 million years to evolve. No primate species pairs have evolved postzygotic isolation on the timescale represented by the evolution of Homo. When archaic and modern humans were in contact, they certainly interbred.
OK, but why is this gene introgression? Why couldn't it just have originated in ancient Africans?
The current evidence for introgression comes from the mismatch between the ancient coalescence time for all haplogroups of the microcephalin gene, compared to the very recent selection on the D haplogroup. Now recent selection on an ancient variant could occur within a single population, for example, if the allele was formerly neutral and gained a new advantage with some difference in the genetic background. And an ancient coalescence date would not be unusual in a single population -- several other loci match the 1.7 million years estimated for the microcephalin genealogy.
Two things make this case especially persuasive. First, there is almost no evidence of recombination between the D and non-D haplogroups. If they existed within the same population for 1.7 million years, they should have recombined a lot with each other, and we should see some of those recombinants today. We don't. The best explanation is that the alleles were in different ancient populations, somewhat isolated from each other so that recombination was very rare.
Second, the D haplogroup is common in Europe and Asia, but is very rare in Africa. If it increased under selection from its origin in some ancient African population, then it ought to be most common in Africa now. We might also expect a deeper origin for the D haplogroup in Africa, similar to the structure of many other genetic loci. We observe neither.
Hey, why should this gene be so unique? There's never been any evidence for archaic genes before!
Now, this is clearly where I have let you down, by not blogging about these papers as they have been coming out. What can I say, I have to make a living somehow! If I give away all my research, how can I stay a step ahead?
The most similar locus to microcephalin is the region around MAPT on chromosome 17. Hardy and colleagues (2005) suggested that this locus is a Neandertal introgression. Like microcephalin, the locus has an ancient coalescence (>2 million years), and like microcephalin, an allele is under selection, with its highest current frequency in Europe. Like microcephalin, MAPT is brain-active, with most research centered on its possible role in Alzheimer's and Parkinson's disease. Unlike microcephalin, there are no recombinants between the major (H1 and H2) haplogroups; this is due to a chromosomal inversion between them. Evans and colleagues (2006) note that balancing selection might not be statistically ruled out when there is such an inversion preventing recombination. Still, balancing selection doesn't easily explain the recent positive selection, nor the geographic distribution of variation.
Garrigan et al. (2005b) found evidence for an ancient Asian allele being retained in living Asians. This allele was from a non-coding locus, so it seems unlikely that adaptive introgression is the cause, which might suggest even more widespread genetic survival of archaic DNA. Some loci suggest the survival of archaic lineages within Africa, including another X chromosome noncoding region (Garrigan et al. 2005a) and the dystrophin gene (Zietkiewicz et al. 2003). These would presumably be attributable to partial isolation of Middle Pleistocene African populations, with introgressive gene flow among them.
The widest survey for introgression thus far was by Plagnol and Wall (2006), who conclude that around five percent of human genes show some evidence for introgression from archaic humans. Their statistical test was looking for loci with ancient divergence times and in particular divergent alleles centered in Eurasian (non-African) populations. So this is a kind of estimate under the assumption of relatively great genetic differentiation among archaic human populations.
I'll end with Templeton (e.g., 2005), who found that human autosomal variation supports a broad ancestry of living humans among Eurasian and African archaics, with evidence of genetic dispersals from Africa several times during the Pleistocene. Under this model, intermixture among archaic populations would have been fairly common, at least intermittently. This is the argument that I made with Milford Wolpoff several years ago (Hawks and Wolpoff 2001) -- we just don't see a lot of evidence for genetic differentiation among archaic humans.
This kind of model would imply that genes like microcephalin -- with strong evidence for some isolation of populations -- might be fairly rare. The fact that several of them have now cropped up (the 5 percent estimate from Plagnol and Wall, 2006, being the most informative on this score) means that we have a lot about archaic human population structure yet to discover.
But notice the nature of this uncertainty. We have a difference between substantial introgression among populations structured like hominoid subspecies on one side, and ubiquitous genetic exchanges among populations structured like human races on the other side. Complete replacement is completely out. "Mostly" replacement, or "assimilation" is still in, but with the observation that archaic human genes had substantial evolutionary importance in the adaptation of modern humans.
In other words, we have moved the ball down the field. Time to line up for the next play.
What is all this about microcephalin possibly not being from Neandertals?
Well, the D haplogroup is common in many areas outside of Africa in addition to Europe. So it isn't possible to really specify in what archaic population it may have originated. There is some chance that it may be found in the Neandertal genome sequence, when that becomes available. In fact, that would be the ultimate test for many candidate introgressive alleles.
But there is a good chance that it won't be found in the Neandertal sequence. After all, Neandertals were probably pretty thin on the ground -- especially in Europe. A sampling of their genes would be sort of unlikely to yield a high proportion of archaic alleles that may have survived to the present day. So there is hope that we will find and document such alleles, but the best evidence for many of them may remain their current pattern of variation in living people.
Now, bear with me here. Neandertals were stupid, right? So why would one of their brain genes be advantageous in modern humans?
There are so many possibilities here.
1. Late Neandertals certainly weren't stupid. Consider the Châtelperronian. And the European Mousterian includes basically all the elements that are thought to represent cognitive sophistication in MSA Africans.
2. Neandertal brains were big, and their heat generation requirements means that energetic constraints were very different from other archaic populations. The brain doesn't function in isolation -- its development, growth, and ongoing maintenance depend on metabolic constraints. So Neandertals might easily have had brain development alleles that had different responses to their high-energy lifestyles. Considering that early Upper Paleolithic people had much more effective foraging strategies than Neandertals, high-energy brain development may have had an even greater advantage than it had previously enjoyed.
3. Modern humans are variable in brain morphology and cognition. That variability certainly includes alternative strategies (for example, personality types) that may be maintained by frequency-dependent selection. An archaic population that had particular constraints on its behavioral strategies might have given rise to strategies that worked within the modern human mix. In that context, Neandertals are fairly unique in having a very strong dietary dependence on meat, and their means of hunting was both risky and required cooperation. That adaptation may have led to behavioral strategies that succeeded in modern humans, even as Neandertal anatomies disappeared.
Those are some possibilities we are working on. There are probably many others. The key is that we are looking at the function of some genes that survived, through our reconstruction of the total phenome of a population that no longer survives. We are limited by the evidence, but there are many suggestive hypotheses.
Neandertals went extinct! Their features disappeared in later humans! How can any of their genes have survived?
This is my favorite one to answer, because it invokes the true paradox of introgression. The features that we recognize as Neandertal features, were defined as Neandertal features by virtue of the fact that they are mostly gone! That means that any alleles correlated with Neandertal morphological features were almost certainly selected against, or were at best neutral. That means that those recognizably Neandertal genes are gone!
But here we have a gene that looks to have come from some archaic population. Adaptive introgression occurs when adaptive alleles are selected, and broken apart from their genetic background. So even as many (perhaps most) Neandertal alleles disappeared, some of their alleles began to increase in frequency -- slowly at first, then very rapidly.
Some adaptive introgressions may already have been fixed, particularly in Europe (from Neandertals). Others, like microcephalin, are still growing in frequency. The key is to remember Mendel -- this is not blending inheritance of Neandertal traits, it is the extinction of many alleles and the proliferation of some others.
The reduction in frequency of Neandertal-like morphological traits over time is entirely consistent with this scenario. In fact, it shows the widespread importance of Neandertal-modern matings, which led to the emergence of a modern population with many Neandertal traits. The widespread genetic contact is documented by the distribution of the traits -- with different Neandertal-like traits in different specimens. That kind of contact is most likely to enable adaptive introgression to proceed.
UPDATE (11/8/2006): Fixed some citations.
References:
Evans PD, Mekel-Bobrov N, Vallender EJ, Hudson RR, Lahn BT. 2006. Evidence that the adaptive allele of the brain size gene microcephalin introgressed into Homo sapiens from an archaic Homo lineage. Proc Nat Acad Sci (early edition) DOI link
Garrigan, D., Mobasher, Z., Kingan, S. B., Wilder, J. A., Hammer, M. F. 2005a. Deep haplotype divergence and long-range linkage disequilibrium at Xp21.1 provides evidence that humans descend from a structured ancestral population. Genetics 170:1849-1856.
Garrigan, D., Mobasher, Z., Severson, T., Wilder, J. A., Hammer, M. F. 2005b. Evidence for archaic Asian ancestry on the human X chromosome. Mol. Biol. Evol. 22:189-192. DOI link.
Hardy, J., Pittman, A., Myers, A., Gwinn-Hardy, K., Fung, H. C., de Silva, R., Hutton, M. and Duckworth, J. 2005. Evidence suggesting that Homo neanderthalensis contributed the H2 MAPT haplotype to Homo sapiens. Biochemical Society Transactions 33:582-585.
Hawks, J., Wolpoff, M. H. 2001. The accretion model of Neandertal evolution. Evolution 55:1474-1485.
Plagnol, V., Wall, J. D. 2006. Possible ancestral structure in human populations. PLoS Genet. 2:e105. DOI link.
Templeton AR. 2005. Haplotype trees and modern human origins. Yrbk Phys Anthropol 48:33-59. DOI link
Zietkiewicz, E., Yotova, V., Gehl, D., Wambach, T., Arrieta, I., Batzer, M., Cole, D. E., Hechtman, P., Kaplan, F., Modiano, D., Moisan, J. P., Michalski, R., Labuda, D. 2003. Haplotypes in the dystrophin DNA segment point to a mosaic origin of modern human diversity. Am. J. hum. Genet. 73:994-1015.
Why introgression?
I heard from a long-time correspondent this morning concerning introgression of microcephalin from archaic humans. I'm not sharing the whole message, but I thought it would be worth paraphrasing a key point for some thought.
The basic point is this: Why are we talking about "introgression"? Why isn't this just gene flow?
Let me start by saying this: "Introgression" is a useful term because it conveys a genetic reality, regardless of the taxonomic rank we are talking about. The literal meaning is "moving into", and what we are talking about is an allele moving into a new population. But more than that (and what distinguishes the term from gene flow) we are talking about an allele moving onto a new genetic background. The "genetic background" implies that there might be constraints on the movement of such an allele coming from epistasis or negative effects of linked alleles.
I think it is especially useful in the case of MCPH1 because we are interested in the clear positive selection of this allele as a contrast to the clear decline in frequency of most archaic morphologies. The differential fates of different genes seem like a good example of some genes introgressing into a new genetic background.
Now, one may object that "genetic background" isn't really a meaningful term. At the very least, it isn't very specific -- it might be better to have a list of genes that interact with each other and exert epistasis on potential introgressions. But it has the virtue of being empirically quantifiable. The overall genetic differences between archaic humans will eventually be measured, including their differentiation from the later modern population. As I mentioned in the FAQ, we can't narrow these values down right now, but more knowledge of genomics is going to make it quite possible. I think that the idea of archaic genes moving into a modern genetic background is going to describe the some of the evolution of early modern humans -- and I think these are important because they are selected. In other words, it is their dynamics that makes them important, not the other way around.
UPDATE (11/9/2006): Razib gets into the introgression-defining act:
Gene flow is a generic term, and can correctly characterize a whole host of dynamics, while introgression is very specific and precise, a subset of gene flow rather than a synonym.
The description that follows is worthwhile, but it is a little problematic. For instance, there is the introduction of a hybrid zone as a mediator through which introgressive genes move in the process of transfer from one population to another. From some points of view this seems to work. For example, cottonwoods in Utah have well-defined hybrid zones (determined by altitude), through which introgressive alleles are thought to have passed, although now they are distributed widely into the range of the opposite parental population.
But lots of other populations don't have hybrid zones at all. Wolves and coyotes (and dogs) mate fairly extensively wherever they are sympatric. Bison had a time in history when they received lots of genes from cattle, and introgression has continued here and there. There was never any well-defined hybrid zone, unless we consider the entire surviving population of bison to have been the zone. Introgression from mountain hare into European hare in Spain seems to have been structured around ancient Pleistocene contact zones rather than current distributions.
And the whole concept of a "hybrid zone" doesn't really apply well to subspecific interactions.
More in my new post, "What about species?"
Look to the baboons; there will you your insights find!
Clifford Jolly's review article in the 2001 Yearbook of Physical Anthropology pretty much covers every aspect for which baboons make an analogy for human evolution. These include Jolly's own "seed-eaters" hypothesis, the implications of baboon diversity for early hominid diversification, the spread of features across geographically dispersed populations, and the implications of baboon hybrids for hypotheses of modern human origins.
Jolly spends a lot of time talking about the implications of hybrid sense and population replacement for the evolution of early Homo. It's interesting in its new thinking, and well worth going over. There's no sense reviewing it all, since you could just read it, but I'm working on modern human origins problems myself right now and found the following passages relevant:
The fragments of Neandertal mtDNA sequence (Krings et al., 1997; Hoss, 2000; Ovchinnikov et al., 2000) suggest the point at which the Neandertal story can be linked to the analogous history of baboons. Discussion of the Neandertal mtDNA sequence has focused mainly on its relatively ancient separation from the root of all extant human sequences, and its implications for a Neandertal genetic contribution to modern human populations. From the baboon (or chimpanzee, or gorilla) perspective, however, the separation is not very ancient. It is comparable to 600 ka divergences between olive and hamadryas baboon mtDNA haplotypes, and much more recent than, e.g., the Guinea-hamadryas split. Mitochondrial diversity in Papio may be analogous to the condition in Homo before the "event" (generally interpreted as an "out-of-Africa" expansion of a relatively small subpopulation) that eliminated most of the diversity from its collective mitochondrial (and Y-linked, and autosomal) gene-pool. Unfortunately, investigation of continent-wide genetic phenostructure in Papio is still in its earliest stages, so we cannot pursue the analogy further in this direction. We can, however, make some suggestions based on work in contemporary zones of hybridization, especially the Awash anubis-hamadryas hybrid zone. For example, we can conclude that unless an undocumented, radical genetic event occurred in the 600 ka since they shared mtDNA ancestry with the Neandertals, premodern humans were certainly able to interbreed with them and produce viable, fertile, offspring, as hamadryas and anubis baboons do (Jolly 2001:198).
Jolly notes that evidence of hybrids may only occur within a hybrid zone itself, which suggests difficulty in examining the existence of such scenarios in fossil contexts.
For the human case, this has an important implication: demonstrating phenotypic distinctness (lack of overlap) of Neandertal
and "modern" samples drawn from areas remote in time and space from the zone of contact does not disprove the occurrence of interbreeding at the interface. It also means that the Lagar Velho child, if indeed it is a hybrid, is a rare and valuable find, even though it is irrelevant to the Neandertal "species question," and does not tell us whether Neandertals (or other "archaic" humans) contributed genes to the Upper Paleolithic, or the extant, human gene-pool. Not that these are equivalent, as is often implied; there was ample opportunity for the loss of a few stray Neandertal genes from European Upper Paleolithic populations when the latter shrank and were replaced by food-producing peoples (Jolly 2001:198-199).
This is probably true from a morphological point of view -- morphological mixture will be evident only at the time and place where different populations were clearly in contact. It is less true of individual alleles or features, which might well intergrade much more extensively depending on their selective dynamics. The least persistent evidence of mixing will be features that are substantially multigenic -- which of course probably includes most of the anatomical features with which anthropologists are familiar. On the other hand, individual features or genes might well be expected to persist long after Neandertals themselves disappeared. This appears to be the case for the features examined by Frayer (1993). In that case, the finding of actual "hybrid" individuals is not so relevant: what is important is the observation of changes in trait frequency within larger, more temporally-dispersed samples.
Jolly later discusses the dynamics of hybrid zones and their application to the discovery of Neandertal-modern mixture:
In the Neandertal case, the fact that the interface moved historically from east to west indicates that the pressure of gene-flow was greater in that direction; if a hybrid zone existed, the genes in it were contributed disproportionately by "moderns." "Neandertal morphological genes" may have been removed by natural selection from a narrow zone of hybridization, or been swamped by differential genetic inflow, or perhaps they simply died out with their carriers without any hybridization at all. Any combination of these factors could have contributed to their disappearance. A much more fine-grained temporal record of the transition would be necessary to decide between these alternatives, and the precise scenario is immaterial both for the eventual outcome, and for the so-called "species question" (Jolly 2001:199)
What is important, and hotly contested, is whether Neandertals (and other archaics) contributed any genes to the gene-pool of the human population who succeeded them. This would imply a
flow of genes from the marginal hybrid zone into the expanding modern population: swimming, as it were, against the tide. The important question is not whether Neandertals could have passed some genes by hybridization to incoming Afro-Arabians; they almost certainly could. It is certainly not the neoessentialist (Cartmill, personal communication) red herring of whether or not they were "really" different species. The important questions are purely empirical: first, whether they actually did contribute any distinctive alleles to the incoming population, and second, whether any of these have survived post-Pleistocene upheavals in the human gene-pool. The first question can only be answered by genetic investigation of the DNA of post-Neandertal fossil humans (cf. Hawks and Wolpoff, 2001); the second by trawling the extant human gene-pool itself (ibid.).
Of course, the question of whether they were "really" different species may be deprecated by those who deal with the fuzzy boundaries between species in nature, but "species" is a term that carries loaded meaning for most biologists. Calling Neandertals a different species is tantamount to asserting their irrelevance to the ancestry of recent humans. This is the point behind the "assimilation" model of human origins -- modern humans actually were a different "thing" than Neandertals, and when the two "came into contact," one group "assimilated" the other. If "assimilation" didn't carry this meaning, there would be no reason to talk about it as a model separate from multiregional evolution, or restricted gene flow and isolation by distance, or "mostly Out of Africa", or any number of other names. All these models agree on the presence of Neandertal genes in later people. Where they disagree is in the emphasis. None of them disagree that Neandertals had an evolutionary history different from other regions. But they disagree about just what kind of history that was. So the question of "species" is a central one, not one that can be shoved under a rug.
From that perspective, the important questions are not merely empirical. They are also conceptual. The "assimilation" model depends on a rather complex conceptual scenario. It envisions the differentiation of Neandertal (along with other archaics) and modern populations over some substantial time. During this time, the evolving modern population within Africa gathered steam for its ultimate dispersal, while the Neandertals and other archaics proceeded along their own unique evolutionary trajectories. Finally, the reestablishment of contact among these populations led to the genetic assimilation of most archaic groups and the establishment of a majority-African gene pool throughout the world. In this hypothesis, the initial isolation (which may have been partial or complete) is essential to the ultimate result. Only if Neandertals had become relatively isolated and divergent could their ultimate assimilation make any sense. Thus, the conceptual basis of Neandertal assimilation is their initial speciation, or if not "speciation" in the formal sense, at least their origin as a distinct population and divergence through substantial isolation.
Now this scenario may or may not have happened; we really don't have the data to test it in comparison to its less conceptually elaborate alternatives. But it is not mindless essentialism to note that this hypothesis depends for its reality on a certain historical identity for Neandertals. This identity is potentially testable. Some may find it distasteful to argue about what a species is, since the concept is so variable and messy in its application to living populations, let alone fossil ones. But that doesn't allow us to avoid the issue: what we call things has meaning, particularly to those outside the intricate details of the fossil record.
Jolly points to the baboon example as having important implications for this case of modern human origins. I agree. However, pointing out the analogy between baboon hybrid zones and the Neandertal-modern transition does not make the latter a case of the former. I have no doubt that Jolly would agree quite fully, but it is worth pointing out nonetheless.
In the meantime, the baboons do give a clear notion of the direction that we should look for evidence of Neandertal-modern interactions:
So far, almost all genetic systems investigated in extant humans show no signs of a Neandertal inheritance, but perhaps we need to be more selective in our search. A moving hybrid zone may leave in its wake a few neutral markers derived from the retreating population (Arntzen and Wallis, 1991), but these are likely later to be eliminated by drift. Most likely to survive and be incorporated are genes for traits strongly favored by local conditions (and "hitch-hiking" markers linked to these). Some years ago, a popular work (Kurtén, 1971) plausibly suggested that Neandertals were blond and blue-eyed in adaptation to cloudy, periglacial Europe, while incoming "moderns" had the darker pigmentation of a subtropical people. Perhaps we should survey nordic Europeans for unusually "deep" diversity in noncoding genetic elements closely linked to loci determining pigmentation... (ibid.).
Of course by the publication of this review, precisely that had been attempted by the survey of MC1R variation by Rosalind Harding and colleagues (2000), finding that the so-called ginger allele may be ancient enough to have come from European Neandertals (reviewed here).
Jolly continues with an interesting hypothesis about possible immunological retentions from Neandertals:
Less fancifully, Parham et al. (1994; and Parham, personal communication) speculatively identified a possible Neandertal legacy: an allele of the human MHC system that is found at low frequency in the old Neandertal range. It is remarkable for its inferred ancient separation from other alleles, which themselves form a tight, young clade. MHC alleles are among the likeliest genes to pass through a semipermeable hybrid zone, since selection favors immunological diversity per se, so if the interpretation is confirmed it would set a likely upper limit on the Neandertal genetic contribution to extant Europeans (ibid.).
Much of interest here from the perspective of interbreeding among archaic human groups. Immensely important stuff, and the earlier parts fo the article are just as essential. Please read it.
On a side-note, I also found this interesting mis-citation of myself:
...the uniquely human, culture-driven, in situ conversion of Neandertals to "moderns" (Hawks and Wolpoff 2001) without any appreciable population movement or "gene-flow" is now hard to reconcile with the rather short timescale of replacement (Churchill and Smith 2001), and has been abandoned by its original formulators (Jolly 2001:198).
Funny, I do remember myself ever writing about the "in situ conversion" of Neandertals without gene flow. Nor do I often see gene flow written with scare quotes, since it is given the chapter in most genetics textbooks. Oh well, the idea that people sneakily abandon their theory is seems to be quite the growing meme lately. I guess the only way to avoid it is to keep oneself from "originally formulating" anything.
References:
Jolly CJ. 2001. A proper study for mankind: analogies from the papionin monkeys and their implications for human evolution. Yrbk Phys Anthropol 44:177-204.
As far as cladistics can take mtDNA analysis
In the early access online edition of Genetics, there is a new paper by Toomas Kivisild and (many) colleagues, titled "The role of selection in the evolution of human mitochondrial genomes" (via Dienekes).
The conclusion of the paper is that the appearance of many nonsynonymous mtDNA changes in certain populations may be the consequence of hotspots where mutations happen repeatedly. The rapid mutation rate at these hotspots means that they saturate more quickly than other sites, and their variation in recently-founded populations is therefore higher than expected compared to their variation in more ancient populations. They suggest that the appearance of many non-synonymous variants in "Arctic" populations (found by Ruiz-Pesini 2004) should be explained by the recent colonization of these regions, as opposed to new adaptations to cold in these populations.
The study was a phylogenetic analysis of human mtDNA variation, from a sample of 277 individuals. After deriving a most parsimonious tree, they looked for sites that underwent recurrent mutations in different branches of the phylogeny. These "hotspots" make up a disproportionately large number of the changes within and between human mtDNA lineages. Thus, it is likely that the high proportion of nonsynonymous changes in certain populations might be due to these hotspots.
Within-human coding variation
So does it matter whether or not some human population has a higher number of nonsynonymous variants? If a population did have a higher proportion of nonsynonymous variants, would that be a good sign of local selection?
I would suspect the answer to both questions is no. It certainly makes sense to me, as Kivisild et al. (2005) claim, that the excess of nonsynonymous changes in some populations may be an overrepresentation of nonsynonymous hotspots compared to more limited variation at other sites. So there is a statistical reason besides selection for this observation.
But considering the low global variation of human mtDNA, there shouldn't have been too much opportunity for different regions to become very different in their mtDNA variants. All of them have a recent common mtDNA ancestor, so locally adaptive variants probably don't differ by a large number of substitutions. And if they don't, then we shouldn't expect to see a significant increase in the proportion of nonsynonymous substitutions for those locally adaptive variants. So this is just not a very good test for local selection.
But there is a pretty good test for whether a variant might be a target of selection: Look at its functional consequences. And we now know that many of the variants that are common in different parts of the world actually have functional consequences on life history, degenerative disease, metabolic efficiency, and high-energy tissues like the brain. Some variants are associated with higher cancer rates, some with higher Alzheimer's and Parkinson's rates, some with higher lifespan, and others with greater energy conversion. When these variants differ significantly in their frequencies in different regions, it is reasonable to suggest that they were selected.
Of course testing the hypothesis of selection depends on demonstrating a fitness advantage for the variants, so it remains at least theoretically possible that different individuals have mtDNA with higher or lower cancer risk, lifespan, and energy efficiency without any difference in fitness.
But I don't think that we would make that assumption for any other gene -- it would be silly. And we don't need to know the proportion of nonsynonymous mutations to make that judgement; we just need to know that the gene does something differently in different places.
So I think the paper goes about as far as anybody can in demonstrating the rates of different kinds of mutations from phylogenetic comparisons. But that still doesn't tell us what we want to know: do the genes do anything differently in different populations. And in fact we already know that they do. The phylogenetic comparisons might inform us about how many selected changes there have been since the mtDNA coalescent, but in fact that number must be small because the coalescent is recent.
Comparison of different primate species
This comparison is discussed to some extent in the paper, but it does not become one of the major foci of the conclusion. I think there is more interesting stuff to be found here, and it points to the possibility of significant adaptive evolution in mtDNA sequences across primates.
You might not get this from the conclusion, which suggests that there is little evidence of positive selection in hominoids on the coding regions of the mtDNA as a whole. But read the criteria:
In these tests, maximum likelihood ratios of non-synonymous to synonymous mutations (omega) exceeding 1 are consistent with the hypothesis of positive selection, while values close to 1 indicate selective neutrality, and values converging on 0 suggest strong purifying selection. We conducted both lineage and site specific tests. For the lineage-specific tests, we used a model in which all lineages have the same omega (hereafter referred to as M0) and compared that with a model in which omega is estimated for each lineage (hereafter referred to as M1). To test for the action of selection among amino acid sites within a specific lineage, we compared a model that allows for heterogeneity in omega among sites, but not among lineages, with a model that allows for variation in omega along a predefined lineage (as in (YANG and NIELSEN 2002)) (Kivisild et al. 2005:8).
Negative selection reduces the number of amino-acid coding substitutions (nonsynonymous subtitutions) compared to synonymous substitutions. Positive selection increases it. This test assumes that either negative selection or positive selection has happened, but not both. Of course, there's no easy test to tell whether both might have happened. They alter the ratio of NS/S subsitutions in opposite directions, so the actual NS/S ratio must reflect their force relative to each other. The paper recognizes this problem (p. 18), but doesn't explore it. Is it credible to think that a site that evolves by positive selection in some lineages is not constrained by negative selection in others? If evolution involves the occasional positive selection of variants at sites usually under negative selection, then the test of selection used here will be extraordinarily weak. Indeed, it is significantly stacked against detecting positive selection.
Even so, the phylogenetic comparison of hominoid ( + macaque) mtDNA found that the model that incorporated positive selection at some sites was superior to neutral or purely negatively selected alternatives. Based on this model, the study found that 16 amino-acid codons in hominoids were significantly likely (i.e. p > .95) to have been under positive selection. That seems to me like a bare minimum, as there must probably have been positively selected sites in individual lineages that wouldn't show up in the cross-hominoid comparison.
The total possibility for positive selection on the human lineage seems large. The study found 167 amino acid substitutions separating humans and chimpanzees, compared to 452 between chimpanzees and orangutans (and only 96 between cats and dogs, which seems incredibly low to me). They tabulate the proportion of substitutions from one amino acid to another (e.g. Ala <> Thr, Ile <> Val, etc.), and find that these proportions differ in some cases from the proportion of segregating variants within humans.
Suppose we assume that those 84 of those 167 mutations are human-specific (the paper doesn't include this information). If six of those were positively selected, that's one per million years. If twelve, one per 500,000 years. And there's no reason to think that some of these might not have undergone multiple substitutions; indeed the presence of hotspots suggests that some sites might have been recurrently selected as the genetic background at other sites changed. And it seems likely that the 414 amino acid segregating variants in humans might include some that had been selected previously during human evolution also. How many selected substitutions may have happened during recent evolution cannot yet be estimated, but how surprising should it be that the most recent one happened around 160,000 years ago?
An aside
Here's an interesting suggestion; I wonder if it's true:
One factor that could, theoretically at least, explain the different amino acid replacement patterns observed between populations and between humans and other mammals is diet. Threonine and valine, essential amino acids that must be taken in the diet, are abundant in meats, fish, peanuts, lentils, and cottage cheese, but deficient in most grains (Kivisild et al. 2005:17-18).
It's another possible reason for selection based on diet during the last 10,000 years. If it affects metabolism strongly enough, which remains to be demonstrated.
Do I have to keep writing about mtDNA?
I'm sure some readers are beginning to think this is mtDNA Selection Central. Believe it or not, I've gotten a lot of requests to cover this topic, which of course is one of the central issues in the Neandertal problem as well as the unraveling of human origins.
And it's an exciting developing story: it shows how medical genetics is steamrolling the human genetics of the past thirty years. Finding mutations that actually do things has great medical interest, and the search is accelerating. This work is being undertaken by people who have no investment in the idea that variation among humans should be completely neutral.
After all, what's more important: that a neutral mtDNA lets us trace human migrations, or that understanding mtDNA selection helps us find treatments for Alzheimer's disease? There's no way that obsolete lineage tracing can survive this kind of conflict. Finding out the history of mtDNA variability is telling us something very important, but it isn't about the movements of people around the globe 100,000 years ago. It's about the evolutionary tradeoffs that led to advantages and disadvantages for different variants.
References:
Kivisild T et al. 2005. The role of selection in the evolution of human mitochondrial genomes. Genetics (online before print).
Neandertal mtDNA from El Sidrón, Spain
Lalueza-Fox and colleagues (2005) report on the recovery of endogenous mtDNA from a Neandertal specimen from El Sidrón cave, in northern Spain. The fossil specimen (El Sidrón 441) was dated by AMS to a calibrated 43,129 +/- 129 years. Only a very limited amount of sequence could be recovered, amounting to only two short fragments of under a hundred base pairs total. This area of HVR-1 contains a number of distinctive mutations in other Neandertal sequences, however, and the El Sidrón reconstructed sequence is identical to the Feldhofer 1 (but not Feldhofer 2) and the two Vindija sequences over at least its short length.
The authors push two interpretations of this variation. The first focuses on the fact that the Spanish specimen shares its sequence with the Vindija specimens of a similar age. These individuals had the length of Europe between them, and may have belonged to populations that inhabited different refugia during glaciations. Taking these assumptions, the authors conclude that it would be unlikely for two distant specimens to share a sequence unless the effective size of the European population were very small. I would say this speculation is rather far from the data, considering that only a very short length of sequence is available, and that the known Neandertals vary at only one nucleotide in this short length.
The second interpretation concerns the time period over which the known Neandertal diversity accumulated. The authors seem to be concerned with the hypothesis that Neandertal genetic variation may have been greatly limited by the glacial maximum that occurred around 130,000 years ago. They estimate the dates at which mutations happened among the known Neandertal sequences, under several different assumptions about which changes were important or should be neglected. The most relevant, excluding the probable artifacts of the Feldhofer 1 sequence but including the long sequences available to date, estimates 162,000 ± 41,700 years for the most recent common ancestor and 92,000 ± 25,000 years for the mutation at site 16,258 shared by the El Sidrón sequence and the Feldhofer 1 and Vindija sequences. I don't thing there's really any story here, although if these dates are accurate, they show that the Neandertal mtDNA substantially postdates the origins of most Neandertal features in the Middle Pleistocene European sample. In this context, the interesting question is what factors led Neandertal variation to be comparably limited compared to human variation.
References:
Lalueza-Fox C et al. 2005. Neandertal evolutionary genetics: mitochondrial DNA data from the Iberian peninsula. Mol Biol Evol Advance Access.
Would you eat them in a box?
Rex Dalton's piece on Neandertal DNA in last week's Nature included this:
The DNA bug has also bitten [Alban] Defleur, who is seeking to collaborate with one of the gene-hunting teams. His group has already found dozens of fragments of Neanderthal bones at Moula-Guercy, representing at least six individuals, both juvenile and adult. "I think there are many more here," says Defleur. [Tim] White, a palaeoanthropologist at the University of California at Berkeley, has excavated with Defleur from France to Ethiopia and says: "Defleur's high-quality excavation and recovery at Moula-Guercy rivals that of modern forensics."
Sounds like a push is on -- will field sites get more funding if they have the promise of DNA retrieval? Will this draw more money away from sites without great faunal preservation (from which one might infer the possibility of hominid bone preservation)?
A key advantage of the Moula-Guercy site is that the bones are well preserved -- because of the cannibalism that removes the flesh and thus destructive bacteria. Long bones have been cracked open for the marrow, which may increase the likelihood that DNA will survive, says White, an authority on cannibalistic practices.
White suspects this may be why Pääbo was successful in sequencing DNA from the Croatia samples: because they were from a site, called Vindija, where cannibalism was practised.
Hmm...
Diet and mtDNA selection
Lowell and Shulman (2005) report on the possible links between the metabolic defects underlying type 2 diabetes and mitochondrial dysfunction. These links go through two channels. In the first, decreases in mitochondrial activity in older adults were associated with higher levels of triglycerides in muscle and liver tissue as well as greater insulin resistance in muscle tissues. This observation supports the hypothesis that mitochondrial oxidation of fatty acids becomes less effective in older individuals, "which in turn lead[s] to increases in intracellular fatty acid metabolites...that disrupt insulin signalling'' (384). It is not clear whether this alteration is due to mitochondrial loss or reduction in function, but the authors suggest based on several other studies that there may be a connection with an accumulation of mtDNA mutations in elderly individuals.
The second channel involves the secretion of insulin by beta cells in the pancreas. In individuals with insulin resistance, the body can sometimes adapt to greater insulin requirements by ramping up the production of insulin in the pancreas. This pathway of insulin secretion depends on the mitochondrial metabolism of the beta cells. This connection has been established by the fact that mtDNA mutations can induce hereditary diabetes by causing beta cell dysfunctions.
Changes in fatty acid metabolism would likely be necessary at least twice during the evolution of early humans. With a dietary change toward greater meat eating, either at the origin of the habilines or that of early large-bodied Homo, a greater dietary availability of animal fats and focus on those resources might well have driven a selective change in digestive metabolism. The highly meat-dependent diet of people in the northern extremes, including the Neandertals, would have focused most digestive and metabolic resources toward animal protein and fat, and might have required additional changes. Then, a shift from a Neandertal-like diet to a broader diet during the Upper Paleolithic might well have required an additional change. It is not obvious that these shifts occurred globally, and there may well have been regional differences in meat digestion and metabolism based on local selection due to dietary differences. If the mtDNA was one of the genetic regions affected by such selection, there may well have been a very complex pattern of evolutionary changes in this molecule across the human lineage. This could account for changes within the Neandertal lineage, as well as the apparent replacement of Neandertal mtDNA by a type prevalent in recent humans.
References:
Lowell BB and Shulman GI. 2005. Mitochondrial dysfunction and Type 2 diabetes. Science 307:384-387.
Max-Planck Institute aims to build Neandertal genome
It's a short story from the AP, but here's the lede:
FRANKFURT, Germany - German and U.S. scientists have launched a project to reconstruct the Neanderthal genome, the Max-Planck Institute for Evolutionary Anthropology said Wednesday.
The project, which involves isolating genetic fragments from fossils of the prehistoric beings who originally inhabited Europe, is being carried out at the Leipzig-based institute.
There are no details.
Now, what is the real story here? I suggest some points to consider:
- The head of Max-Planck Evolutionary Anthropology is Svante Pääbo, who is the world's leading expert on DNA recovery from ancient remains. In my assessment, he is very unlikely to announce a project like this one without some advance knowledge that there will be results.
- The recovery of a complete mitochondrial genome from Neandertals is certainly achievable. But nobody from Max-Planck or anywhere else has yet published such a sequence. It would also be actually informative, in that it would give us some information about the amino acid sequences coded by human mtDNA prior to the common ancestor of all living human mtDNA. This would be the opportunity to see which evolutionary changes in mtDNA might have been targets of recent selection.
- But, there would be no reason for a press release about a complete mtDNA genome in the absence of details. Anybody can see that a complete mtDNA sequence is achievable, and while it would be important news, it would not justify this kind of attention.
- Therefore, they really are talking about recovering nuclear DNA information from Neandertals.
- Referring to point (1) above, this means that someone in Pääbo's lab has very likely already recovered some nuclear sequence from a Neandertal.
- That lab has, in recent publications, focused at some length on the problem with contamination inherent in Neandertal remains, and has been skeptical of results from other labs reporting modern human sequences from post-Neandertal modern Europeans. The clear trend over the past two years is to reject any modern sequence as possible contamination.
- Using this standard, we may conclude that at least some nuclear DNA thus far recovered from some Neandertal specimen exhibits some interesting difference from living humans.
Much of this train of logic follows directly from point (1), that Pääbo is unlikely to make an announcement like this without already knowing some results. This assumption is pretty justifiable for anybody familiar with the literature. After all, Pääbo has himself been highly critical of botched ancient DNA recoveries by other labs. Also, the very existence of news stories suggests strongly that a negative result is not the end point.
If you have any lingering doubts, then read this 2003 story from US News and World Report, where Richard Klein says "Svante is going to be the first anthropologist to win a Nobel Prize." I think my assessment is very likely; this is not someone who is going to be associated with a highly publicized failed effort.
Assuming that some Neandertal nuclear DNA is coming, what would be useful? First of all, the stated goal of "reconstructing the Neandertal genome" is completely impossible, unless by this they mean the mtDNA genome. The focus will be on highly polymorphic sites that contain lots of single nucleotide (or other short) polymorphisms. This is a good reason to expect there will not be Y chromosome evidence, despite its probable interest, because the Y is just not very polymorphic. Longer length polymorphisms, like microsatellites, likewise are not very realistic because they would be very difficult to clone without error from a damaged sequence, and they extend over dozens to hundreds of base pairs, making it difficult to find them (along with enough flanking sequence for primers) intact. It is possible that they will focus on the X chromosome, which is considerably easier to deal with in males because they do not carry two potentially different copies.
The twist is that you are not going to confirm the DNA is real (and not contaminated) unless you can show a difference from living humans. My gut tells me that for most nuclear genes those kinds of differences are unlikely to be found. A real advance in studying these ancient remains would be some theoretical movement on the likelihood of observing certain kinds of presently rare alleles in the same individual. If I were doing this work, I would work out the odds of observing certain combinations of biallelic sites in present-day Europeans, and then assess the same sites in a Neandertal fossil. This would allow a more-or-less standard admixture study, albeit with a very small ancestral sample. Or, I would try to find specific short functional alleles that are polymorphic in humans. For example, MC1R would be very interesting, because you could try to find haplotypes selected for skin or hair color. This story would be about the antiquity of selection for skin color, although it might not necessarily demonstrate a Neandertal origin for light skin in Europeans.
But I don't think they'll do what I would do.
My best guess is that we will see first the sequence for FoxP2 from one or a few Neandertal specimens.
Pääbo's lab has done much of the molecular work on FoxP2 (OMIM), including assessment of the variation in other primates. This means they have primers for the gene that work both within humans and among different primate species. Furthermore, the gene underwent a recent selective sweep (dating to the last 200,000 years) in humans, meaning that Neandertals are fairly likely to lack the present human allele. Humans differ from chimpanzees by two amino acid sites, so it is likely that one of these differs between recent humans and their ancestors. Although it is possible that Neandertals are identical to humans, there is still good reason to suspect that Neandertal FoxP2 sequences will be different. And such a difference would help pass the contamination test.
It's like the perfect genetic storm.
You can expect that if this is the finding, the conclusion will be a speculation about Neandertal speech abilities. I have recently written with some colleagues (Wolpoff et al. 2004) about this issue, concluding that FoxP2 just doesn't tell us much about speech and language in Neandertals. So it will be interesting to see anthropology and genomics interacting if we get more information about Neandertal genetic variation.
To you graduate students in Leipzig, please watch out! I know that many archaeologists think Neandertals were dumb. But you can bet that if the velociraptors could learn to open doorknobs, then cloned Neandertals can too.
UPDATE (7/7/05): A reader has sent me a reference to a paper in press describing nuclear DNA recovery from cave bear remains. I discuss it in a later post. That success may explain the new confidence about the prospect for Neandertal DNA, but I'm still thinking there probably is already some positive result from them also. The scope of the bear research is very suggestive about the method, however, and it is much more akin to what I think would be informative than I feared.
References:
Wolpoff MH et al. 2004. Why not the Neandertals? World Archaeol 36:527-546. PDF available online
Neandertal genome project
By now you've probably read something about the Neandertal genome project. But have you seen the press kit from 454 Life Sciences?
Due to such sample contamination, the task of sequencing the Neandertal genome is much more extensive than the task of sequencing the human genome. 454 Life Sciences' Genome Sequencer 20 System makes such an endeavor feasible by allowing approximately a quarter of a million single DNA strands from small amounts of bone to be sequenced in only about five hours by a single machine. The DNA sequences determined by the Genome Sequencer 20 System are 100-200 base pairs in length, which coincides neatly with the length of ancient DNA fragments.
Of course, there's nothing really new here; they're just saying what their goals for the project are. The only concrete result is this:
Approximately 99% of the Homo sapiens genome is identical to the chimpanzee genome, our closest living relative. It is estimated that the Neandertal shares 96% of the 1% difference with Homo sapiens. The Neandertal shares the remaining 4% of the difference with the chimpanzee.
That has been part of the public talks about the sequencing, as well as the press conference. If we assumed a 7 million year genetic divergence of humans and chimpanzees, this would place the human-Neandertal genetic divergence time at 560,000 years ago. They don't mention the obvious: most human genes have variation that is a lot older than 560,000 years -- so Neandertals will be within the human range of variation for most genes.
The hope of the project is to identify the set of genes that were under selection too long ago to detect them in recent selection assays, but more recently than the average human-Neandertal genetic divergence. In other words, genes for which the Neandertal allele no longer exists in living people.
But there's another use we might consider, raised by Bruce Lahn in a New York Times article by Nicholas Wade:
A longstanding dispute among archaeologists is whether the modern humans who first entered Europe 45,000 years ago, ultimately from Africa, interbred with the Neanderthals or forced them into extinction. Interbreeding could have been genetically advantageous to the incoming humans, says Bruce Lahn, a geneticist at the University of Chicago, because the Neanderthals were well adapted to the cold European climate -- the last ice age had another 35,000 years to run -- and to local diseases.
Evidence from the human genome suggests some interbreeding with an archaic species, Dr. Lahn said, which could have been Neanderthals or other early humans.
Of course, the Neandertal genome would be very important for confirming possible genetic contributions from Neandertals.
I think the most interesting thing will be the variation in Neandertal genes. For that question, I think the project is facing a pretty extreme challenge:
Over the next two years, the Neandertal sequencing team will reconstruct a draft of the 3 billion bases that made up the genome of Neandertals. For their work, they will use samples from several Neandertal individuals, including the type of specimen found in 1856 in Neander Valley and a particularly well-preserved Neandertal from Croatia. The Max Planck Society's decision to fund the project is based on an analysis of approximately one million base pairs of nuclear Neandertal DNA from a 45,000-year-old Croatian fossil, sequenced by 454 Life Sciences.
OK, here's the thing: they have to try to assemble a sequence from a diploid individual. This is really challenging, because anywhere the individual is a heterozygote, there will be ambiguity about which SNPs are linked on one chromosome, and which belong together on the other.
Now, they faced the same problem in the mammoth Mc1r paper, which they resolved by sampling additional mammoth individuals and finding homozygotes. That may be the reason for mentioning the "several Neandertal individuals" in the press release. But it's not obvious that even the "several" they are likely to have samples from will be enough for most genes to be unambiguously reconstructed.
And then there's the contamination problem. Well, that one I'll save for another day.
The "flame-haired" Neandertals
What better to match an Irish tongue, than red hair? In fact, with language and red hair Neandertals would seem to be excellent candidates for a little scheme I have copying the encyclopedia...
Well, that's the story, anyway. How true is it? It's been two weeks since the hea