What killed off Neanderthals around 40,000 years ago has long puzzled scientists.
Some have suggested that as humans migrated from Africa they out-competed Neanderthals for resources, while others claim climate change finished them off.
Now, a new study shows that - whatever it was that drove Neanderthals to extinction - the species was doomed from the start.
A computer simulation showed that the slow but continuing trickle of humans coming from Africa to Eurasia meant they outlasted the Neanderthals.
Even without human competition or a wavering climate, Neanderthals would have died out thanks to random genetic drift, the research found.
The issue of admixture between Neandertals and modern humans has been contentious. The arguments for and against such admixture were raised in several paleontological studies (Tattersall and Schwartz 1999; Klein 2003; Zilhao 2006; Trinkaus 2007). Neandertal mitochondrial DNA sequencing provided no evidence of Neandertal matrilineal contribution to contemporary humans (reviewed by, Hodgson and Disotell 2008). In contrast, a draft sequence of the Neandertal nuclear DNA revealed the presence of a number of bona fide Neandertal segments in non-African genomes (Green et al. 2010). Yet, although rather unlikely given all the precautions taken to eliminate such a possibility (Green et al. 2010), the admixture results could be due to contamination of ancient DNA with modern human genomic fragments (Wall and Kim 2007). In this paper, we provide further evidence of Neandertal admixture. Our laboratory did not participate in the determination of the Neandertal sequence. The admixed X-linked haplotype of Neandertal origin we report was suspected to descend from a non-African lineage well before the advent of the Neandertal genome (Zietkiewicz et al. 2003).
Table 1. Twelve Major dys44 Haplotypes and the Neandertal Sequence.
To the admixed haplotypes from the bulk of genomic diversity, we assume that modern humans left Africa somewhere between 80–50 thousand years ago (Kya) and that at that time Neandertals occupied western Eurasia (Stringer 2002; Klein 2003; Mellars 2006; Oppenheimer 2009; Petraglia et al. 2010). Because these populations diverged 400–800 Kya, a number of derived alleles present in the Neandertal DNA can be expected to segregate in all human populations including sub-Saharan Africans. In contrast, segments in human DNA that were subsequently admixed outside Africa are expected to carry additional derived alleles shared with Neandertals but absent in sub-Saharan Africans. The admixed haplotypes are also expected to be younger, that is, less diversified by recombination than the average African haplotypes at the same loci. Ideally, the allelic structure of these haplotypes would differ from the bulk of the common haplotypes reflecting their origin along a separately evolving lineage.
An X-linked haplotype that fulfills these characteristics occurs in an 8-Kb intronic segment spanning exon 44 of the dystrophin gene, referred to as dys44 (Zietkiewicz et al. 1997). We analyzed dys44 polymorphisms in a sample of 6,092 X-chromosomes representing populations from all habitable continents (supplementary table S1, Supplementary Material online), including previously published data (Labuda, Labuda, et al. 2000; Zietkiewicz et al. 2003; Xiao et al. 2004; Lovell et al. 2005; Bourgeois et al. 2009). We focus on the 12 major dys44 haplotypes that explain 89% of genetic diversity in non-Africans and 67% in sub-Saharan Africans (table 1). Of these, haplotype B006 is structurally distinct; with only four derived alleles, it is the closest to the ancestral one. Common outside Africa and virtually absent in sub-Saharan Africa (fig. 1), B006 was earlier proposed to represent an unknown non-African contribution to the human gene pool (Zietkiewicz et al. 2003). Of 1,420 sub-Saharan chromosomes, only one copy of B006 was observed in Ethiopia, and five in Burkina Faso, one among the Rimaibe and four among the Fulani and Tuareg, nomad-pastoralists known for having contacts with northern populations (supplementary table S1, Supplementary Material online). B006 only occurrence at the northern and northeastern outskirts of sub-Saharan Africa is thus likely to be a result of gene flow from a non-African source.
FIG. 1. Worldwide distribution of B006 haplotype based on a sample of 6,092 X-chromosomes. Samples are listed in supplementary table S1, Supplementary Material online. Certain subpopulations were merged, when justified by their geographic proximity.
In the available Neandertal sequence (Green et al. 2010), there is information on 20 of 35 dys44 polymorphic sites. These represent 18 ancestral and 2 derived alleles, fully matching the corresponding sites of B006 (table 1). One of the derived alleles, C of rs6631517, is also shared with other dys44 haplotypes, whereas the second one, G of rs11795471, is unique to B006 (the information on two remaining B006-polymorphisms is not available). Figure 2 illustrates plausible historical pathways leading to the three observed categories of the dys44 haplotypes. Haplotypes, such as B007, B010, and B012 in table 1 are specific to sub-Saharan Africa. They carry common (sites of type 1 and 3) and African-specific polymorphisms (sites 2, 4, and 6). The remaining haplotypes, except B006, are cosmopolitan and are found both inside and outside Africa. The third category, absent from Africa, is represented by B006, which carries two types of derived alleles that are shared with Neandertal DNA. The mutation at site 1 (as rs6631517 above) was presumably fixed in Neandertals and segregated along the ancestral allele in the human lineage. Derived alleles acquired through recent admixture (site 5) are expected to be only found in the progeny of the admixed individuals as in the case of rs11795471. Moreover, the Neandertal origin of B006 is consistent with the allelic status of the remaining 19 of the 20 Neandertal sites of known identity (table 1) and confirmed by analysis of the B006 flanking segments in HapMap3 (Altshuler et al. 2010).
FIG. 2. Scheme of evolutionary pathways leading to three categories of haplotypes in the dys44 segment. Twelve of the 35 dys44 polymorphisms available in the HapMap3 database are sufficient to identify the 12 major dys44 haplotypes presented in table 1. In the considered HapMap3 populations (supplementary table S1, Supplementary Material online), these haplotypes represent 369 of 523 (70.6%) sub-Saharan chromosomes and 815 of 891 (91.5%) non-African chromosomes that include 77 copies of B006. We extended the dys44 8-kb region by 28 additional polymorphisms on the left and 49 on the right (108 kb in total) to include sites that by inspection appeared to maintain some degree of linkage disequilibrium with the B006 haplotype (fig. 3). In the Neandertal sequence, no information is available for 28 of these sites, 36 sites represent ancestral alleles and 13 derived alleles. Importantly, three of the derived alleles (rs17243319, rs1456729, and rs11796299 in supplementary table S2, Supplementary Material online) are absent from the African chromosomes, as in the case of the derived G of rs11795471 from within B006. Moreover, all derived alleles shared with Neandertals occur at high frequencies (0.75 and more) on a background of the extended B006 haplotype (fig. 3 and supplementary table S2, Supplementary Material online) as expected in a segment of recent Neandertal origin.
Outside Africa, B006 is found in all habitable continents including Australia, as determined from a remote community of isolated indigenous Australians living in Central Australia (fig. 1). The ubiquity of B006 lineage reflects a worldwide contribution of Neandertal lineages to non-African genomes. It indicates very early Neandertal admixture prior to successful range expansion of the population ancestral to virtually all contemporary non-African populations and confirms earlier contention of very early admixture based on the analysis of Neandertal segments in European, Han, and Papuan genomes (Green et al. 2010). Hodgson et al. 2010 proposed such admixture through early Levantine contacts of modern humans and Neandertals, prior to the most recent out-of-Africa expansion, and suggested that traces of such admixture should be still detectable in sub-Saharan populations of Northeastern Africa.
FIG. 3. Derived allele frequencies in the HapMap3-based extended dys44 haplotype. Superimposed histograms compare derived allele frequencies seen on the background of the 77 extended B006 haplotypes (upper) with frequencies of the same alleles on the remaining 1,337 haplotypes from this data set (supplementary table S2, Supplementary Material online). The status of the Neandertal alleles is marked on the top as ancestral, derived, or not available, NA.
In contrast, rather than considering admixture, an alternative explanation posits that the ancestral population of present-day non-Africans was more closely related to Neandertals than the ancestral population of present-day Africans (Green et al. 2010). Indeed, the evidence is accumulating on deep subdivisions within the ancestral population in Africa (Labuda, Zietkiewicz, Yotova 2000; Falush et al. 2003; Cohen et al. 2007; Yotova et al. 2007; Behar et al. 2008; Gunz et al. 2009). As in the case of admixture through Levantine contacts, the latter explanation also implies significant sharing of Neandertal haplotypes between all non-Africans and Northeastern Africans. On the other hand, no evidence for these scenarios is found in our data, whereas the oldest lineages tend to be found in South rather than Northeastern Africa (Behar et al. 2008; Campbell and Tishkoff 2010). Paleontological findings (Shea 2008; Petraglia et al. 2010) point to the occupation of the Levant by Neandertals and early H. sapiens at nonoverlapping time periods making their early contacts unlikely. Interestingly, some of the HapMap3 haplotypes from the segments proposed by Green et al. 2010 and fulfilling our criteria of Neandertal admixture, also turn out in Maasai, where, however, their occurrence can be due to recent back-to-Africa migration (Sikora et al. 2011).
In contrast to other candidate haplotypes, such as these from “ASPM” and “MCPH1” that seem to have failed the test (Green et al. 2010; Lari et al. 2010), the evidence for Neandertal origin of B006 appears very strong. This provides additional verification of the findings by Green et al. (2010) that Neandertals contributed to the genetic makeup of modern human populations outside Africa. Our data indicate that Neandertal admixture occurred very early or prior to their worldwide expansion. Considering such early encounter of H. sapiens with Neandertals, a question may be raised: was this encounter coincidental and without important evolutionary consequences or (either through genetic or cultural exchanges, or both, Premo and Hublin 2009) did it facilitate adaptations to novel environmental conditions that actually contributed to the successful expansion of human migrants from Africa to other continents?
The challenge for methods seeking to identify introgression is to distinguish true introgression from shared ancestral genetic variation (Figure 1). Any two populations will always share some segments of DNA that originated in their common ancestor because both populations descend from the same population and might therefore have inherited some of the same segments of DNA from the ancestral population. For this reason, two DNA segments sampled from different populations may share a most recent common ancestor (MRCA) more recently than two DNA sequences sampled from the same population. The same argument is true for species, and we will in general here not distinguish between species and populations, in part to avoid entering into discussions of species concepts and definitions in hominins.
Genome-wide admixture maps
Two studies have produced Neanderthal genome-wide admixture maps of European and Asian genomes, based on the whole-genome sequence of a female Neanderthal from Siberia4 and present-day human data from the 1000 Genomes Project 66. In one study, the authors employed a CRF model47 (see Box 2) while in the in the other study, the authors relied on the S* statistic to identify introgressed segments23 (Box 1, Supplementary information S3 (box)). While these two studies find notable signatures of negative selection pushing introgressed variants to low frequencies and a depletion of introgressed tracts in functional genomic regions, possibly due to hybrid sterility, the authors of both studies found considerable evidence of adaptive introgression. Several examples are outlined below. Additionally, one of the studies performed a functional enrichment analysis and found that genes involved in keratin filament, sugar metabolism, muscle contraction and oocyte meiosis have been involved in archaic human adaptive introgression47.
BOX 1 Description of the S* statistic
A different genome-wide study (reviewed in the Metabolism section) employed the D statistic locally to detect Neanderthal-like regions at specific genes.67 The authors found that lipid catabolism genes tended to have significantly higher D values in Europeans (supporting a tree clustering Neanderthals with Europeans, to the exclusion of Africans) than the rest of the genome. Additionally, signals of recent positive selection were reported in the same regions with high D in modern Europeans.67
Finally, HMMs identified within present-day human genome data were used to identify putatively introgressed haplotypes in two other studies4,35. These methods identified present-day human haplotypes that are phylogenetically consistent with forming a clade with an archaic individual (Neanderthal or Denisova). The genes contained in the identified regions were not characterized, and tests of selection were not performed in these cases. However, the divergence between the introgressing Neanderthal in Eurasians and the sequenced Neanderthal genome (77–114 kya assuming a mutation rate of 0.5*10−9 per bp year or 38–57 kya assuming a rate of 10−9 per bp per year) was much younger than the divergence between the introgressing Denisovan in the Sahul people (Oceanians, including native Papuans and Australians) and the sequenced Denisovan genome (276–403 kya assuming the slower mutation rate or 138–202 kya assuming the faster rate).4 This may indicate that the Denisovan population was more diverse and/or more structured than the Neanderthal population.
Examining the candidate genes identified from both the genome-wide and single loci studies, we can speculate on the plausible selection pressures and mechanisms underlying examples of adaptive introgression. We organize our discussion of candidate genes below by broad functional categories.
BOX 2 Hidden Markov Model (HMM) and Conditional Random Field (CRF) frameworks
Defense against pathogens
STAT2 is an innate immune gene that is involved in interferon response after viral infection. A recent study6 found a long (130–260 kb) haplotype called ‘N’ that overlaps this gene and that is broadly distributed across Eurasia. The haplotype is absent in sub-Saharan Africa, has a very ancient time to the most recent common ancestor (TMRCA) with other presentday human haplotypes (609 kya, assuming the faster mutation rate) and also has a very recent TMRCA with the Neanderthal genome (78 kya), suggesting it introgressed from this archaic hominin group. Though found throughout Eurasia at a frequency of ~5%, it is present at substantially elevated frequencies in Papuans (~54%). The authors used a neutrality test that controlled for demography to show this difference in frequency was likely due to positive selection acting on the region in Papuans. Though the study focused on STAT2 due to greater availability of sequence data in this gene, the introgressed N haplotype overlaps two other genes which could also have been the targets of selection: ERBB3 (coding for a peptide involved in cell growth and apoptosis68) or ESYT1 (coding for a transmembrane protein with a role in fibroblast differentation69, 70).
The highly polymorphic human leukocyte antigen (HLA) region in chromosome 6 also seems to show signatures of adaptive introgression, though in this case by balancing selection. The fact that the HLA region is highly prone to trans-specific polymorphisms makes it hard to distinguish adaptive introgression from balancing selection for different variants persisting over long time periods71. Various HLA haplotypes have been identified in Eurasians and Melanesians that carry functional variants that likely introgressed from archaic human groups.5 For example, a deeply divergent haplotype (B*73) with strong sequence homogeneity among its carriers is present at high frequencies in west Asia but absent or infrequent in the rest of the world. Simulations show that the global distribution of this haplotype is best explained by a model of introgression from an archaic source in west Asia.5 The Denisovan genome2 carries two variants (C*12:02 and C*15) that are in strong linkage disequilibrium with B*73, although the actual B*73 allele is absent in the Denisovan genome. These variants are also only found at high frequencies in west Asia. A separate analysis revealed a substantial Neanderthal HLA-A ancestry in modern non-African humans, with a wider geographic distribution than B*73 (ref. 5). In summary, unusual levels of LD and high archaic haplotype frequencies at the HLA locus in present-day humans seem to support the hypothesis of adaptive introgression.
Possible archaic extent of HLA-A allele in modern humans.
Another HLA study examined an important amino acid motif in the family of antigen receptors in the HLA region (HLA-DPβ, allele DPB1*0401) that are required for allowing unmatched DRα and DPβ subunits to form a functional complex. The sequence closely matches the Neanderthal genome at the same locus72. However, a phylogenetic analysis of different variants of this motif in modern humans found that the putatively introgressed haplotype coalesces with other common modern human haplotypes before coalescing with the Neanderthal genome73. Furthermore, although the motif variant is present at 68% frequency across Europeans, it is also present in sub-Saharan Africans at 11% frequency. Finally, the divergence time between the haplotype of interest and the Neanderthal genome was estimated to be 2.2 Mya (assuming a mutation rate of 10−9 per bp per year), largely predating the modern human-Neanderthal population split time. Thus, it is more likely that the presence of this haplotype in Europeans is a consequence of ancient population structure in Africa73.
Other regions of the human genome show characteristic signatures of introgression, but the evidence for positive selection is weak or absent. For example, the OAS gene cluster (which helps to inhibit viral replication as part of the innate immune response) has been subject to two separate archaic introgression events: one into Non-Africans from Neanderthals (though the haplotype is absent in Papuans)74 and another one into Papuans from an extremely deep lineage (TMRCA with modern humans = 3.7 million years ago (Mya), assuming a mutation rate of 10−9 per bp per year)75. The haplotype for the latter matches best with the Denisova sequence across 90kb of high LD sequence, and has much higher sequence diversity than observed in other populations75. Although no evidence for positive selection in the region has been found,74, 75 the genetic distribution of the different haplotypes across continents is perhaps a signal of balancing selection74. The deep lineage could belong to another archaic hominin group that may have admixed with both Papuans and Denisovans, or with a common ancestor of the two populations75. In the same direction, but in a different study, Prüfer et al.4 recently found a genome-wide signal of “super-archaic” introgression in the Denisovan genome, likely due to an unsampled hominin group that diverged from modern humans before Denisovans and Neanderthals, and later interbred with Denisovans. However, the specific regions of the Denisovan genome that have super-archaic ancestry have yet to be identified, so it is unknown if the OAS gene cluster is one of them. Finally, evidence for adaptive introgression into Europeans from Neanderthals in the gene OAS2 was more recently confirmed using the CRF model.47
