Here we present selected parts of the interesting paper titled “A Revised Timescale for Human Evolution Based on Ancient Mitochondrial Genomes”, by Qiaomei Fu et al.
Summary
Background: Recent analyses of de novo DNA mutations in modern humans have suggested a nuclear substitution rate that is approximately half that of previous estimates based on fossil calibration. This result has led to suggestions that major events in human evolution occurred far earlier than previously thought.
Results: Here, we use mitochondrial genome sequences from ten securely dated ancient modern humans spanning 40,000 years as calibration points for the mitochondrial clock, thus yielding a direct estimate of the mitochondrial substitution rate. Our clock yields mitochondrial divergence times that are in agreement with earlier estimates based on calibration points derived from either fossils or archaeological material. In particular, our results imply a separation of non-Africans from the most closely related sub-Saharan African mitochondrial DNAs (haplogroup L3) that occurred less than 62–95 kya.
Conclusions: Though single loci like mitochondrial DNA (mtDNA) can only provide biased estimates of population divergence times, they can provide valid upper bounds. Our results exclude most of the older dates for African and non-African population divergences recently suggested by de novo mutation rate estimates in the nuclear genome.
Introduction Differences in DNA sequences correspond to nucleotide substitutions that have accumulated since their split from a most recent common ancestor (MRCA). When the average number of substitutions occurring per unit of time can be determined, the ‘‘molecular clock’’ rate can be estimated. Under the assumption of constant rates of change among lineages, molecular clocks have been used to estimate divergence times between closely related species or between populations.
Fossil evidence has been frequently used to estimate a date for the MRCA of two related groups, thus providing a calibration point for the molecular clock. The sparseness of the fossil record, however, poses limitations on the reliability of such estimates. For example, in human evolution, no fossil has yet been identified to represent the uncontested MRCA for humans and chimpanzees or other closely related primate species. As a consequence, the nuclear and mitochondrial mutation rates for the human lineage have been heavily debated.
Recent analyses of de novo substitutions from genome sequencing of parent and offspring trios allow the direct calculation of nuclear substitution rates per generation. This alternative to the fossil calibration of the human molecular clock is arguably more accurate. Surprisingly, publications using this approach have recently pointed to de novo rates that are about half the value of those previously determined from fossil calibrations. A slower substitution rate has important implications for inferring the timing of key events in human evolution such as our divergence from our common ancestor with chimpanzees, our divergence from Neanderthals and Denisovans, and the migration of modern humans from one region or habitat to another. Taking these new rates into consideration, most date estimates would be pushed back by a factor of two—for example, yielding a West African/non- African split date of 90–130 kya, which is up to 60 kya older than some previous estimates.
Attempts at calculating the human mitochondrial DNA (mtDNA) substitution rate have relied on either estimates derived from fossil calibration or archaeological evidence of
founding migrations; however, reliance on a single calibration date can easily lead to a biased rate estimate.
An alternative approach for obtaining greater precision in measuring substitution rates is through the analysis of genetic data from ancient samples for which reliable radiocarbon dates are available. Ancient humans are well suited to provide calibration points for the human mitochondrial molecular clock: reliable radiocarbon dates are available for many specimens; hence, the number of substitutions that have accumulated among lineages can be directly translated into the number of substitutions per site per year. Branch shortening—the effect of fewer nucleotide substitutions on ancient branches in a phylogenetic tree as compared to modern—is commonly observed in phylogenetic studies of ancient humans. The observed branch shortening reflects the comparatively shorter time since the common ancestor for the ancient human as compared to the present-day individual: a present-day lineage has had more time to accumulate nucleotide changes. If we know the age of the ancient sample (e.g., from a carbon date), we can thus infer the mutation rate necessary to produce the observed degree of branch shortening.
Here we use the complete or nearly complete mitochondrial genomes from ten ancient modern humans for which reliable radiocarbon dates are available to calculate the human mtDNA substitution rate directly.
Evolutionary Analysis All but one of the ancient modern human sequences from Europe belonged to mtDNA hg U, thus confirming previous findings that hg U was the dominant type of mtDNA before the spread of agriculture into Europe.
For the remaining eight ancient Europeans, we built a phylogenetic tree for hg U, which included 63 contemporary mtDNAs from this haplogroup. The tree clearly shows that all
four Paleolithic pre-Last Glacial Maximum (LGM) samples display a short branch compared to the four ancient post-LGM samples. Predictably, the older samples Dolni Vestonice 14 and 15 fall in a basal position relative to the contemporary mtDNA hg U5. The Tianyuan sequence from Eastern China falls basal to the contemporary hg B, common in most parts of Eastern Asia, Oceania, and the Americas. The mtDNA genetic diversity that we measure in early modern Europeans is about 2-fold less than the mtDNA diversity in today’s Europeans, but about 1.5 times higher than that measured in Neanderthals contemporary with these early modern humans (excluding the older Mezmaiskaya individual). Although these measurements provisionally suggest that a higher population size might have contributed to early modern humans outcompeting Neanderthals after their arrival in Europe, there are caveats to this analysis. First, we have a limited sample size of ancient specimens. Second, we have sampled from several different time periods, a practice that overestimates actual genetic diversity. Third, our sampling is nonrandom; for example, we included several individuals from within the same burial site (Dolni Vestonice), where maternal relatedness is possible and would give an underestimate of true diversity. More data are necessary to provide a definitive assessment of the genetic diversity of these prehistoric populations.
Substitution Rate Estimates For the linear regression approach we estimate a substitution rate of 1.9231028 per site and year (1.16–2.6831028, 95% CI) for the whole mtDNA and 1.25 6 0.68 3 1028 per site and year (0.57–1.93 3 1028, 95% CI) for the coding region.
The substitution rates for the mtDNA coding region and whole mtDNA largely overlap with the above results when the four radiocarbon-dated Neanderthals are included alongside the ten ancient modern humans.
In theory, mitochondrial substitution rates could have changed between Neanderthals and modern humans, though we do not detect evidence of this because inclusion of Neanderthal data does not lead to a rejection of the molecular clock. We use only the substitution rates calculated with radiocarbondated ancient modern humans to calculate modern human mtDNA divergence times, given that substitution rates among modern humans are most relevant to estimating divergence times among modern humans and that the Neanderthal data add little extra statistical precision.
Haplogroup Divergence Time Estimates Using the substitution rate for the whole mtDNA genome obtained by Bayesian estimation, we estimated the time of the MRCA for all modern humans at 157 kya (120–197 kya, 95% HPD). Our rate also implies a split of all non-African hgs from the closest widespread sub-Saharan African hg (L3) of 78.3 kya (62.4–94.9 kya). The MRCA of hg Q, often referred to as a maximum age for the settlement of Australia, was calculated at 42 kya (30–54.9 kya). The time to most recent common ancestor (TMRCA) of hg U5, often argued to have evolved within the first early modern humans in Europe, was calculated at 29.6 kya (22.7–37.2 kya).
Discussion We were able to reconstruct three complete and six nearly complete mitochondrial genomes from ancient human remains that were found in Europe and Eastern Asia and span 40,000 years of human history. All Paleolithic and Mesolithic European samples belong to mtDNA hg U, as was previously suggested for pre-Neolithic Europeans. Two of the three individuals from the Dolni Vestonice triple burial associated with the pre-ice age Gravettian culture, namely, 14 and 15, show identical mtDNAs, suggesting a maternal relationship. Furthermore, both individuals display a mitochondrial sequence that falls basal in a phylogenetic tree compared to the post-ice age hunter-gatherer samples from Italy and central Europe, as well as the contemporary mtDNA hg U5. It has been argued that hg U5 is the most ancient subhaplogroup of the U lineage, originating among the first early modern humans in Europe. Our results support this hypothesis because we find that the two Dolni Vestonice individuals radiocarbon dated to 31.5 kya carry a type of mtDNA that is as yet uncharacterized, sits close to the root of hg U, and carries two mutations that are specific to hg U5. With our recalibrated molecular clock, we date the age of the U5 branch to approximately 30 kya, thus predating the LGM. Because the majority of late Paleolithic and Mesolithic mtDNAs analyzed to date fall on one of the branches of U5, our data provide some support for maternal genetic continuity between the pre- and post-ice age European hunter-gatherers from the time of first settlement to the onset of the Neolithic. U4, another hg commonly found in Mesolithic hunter-gatherers, has so far not been sequenced in a Paleolithic individual, and we find hgs U8 and U2 in pre-LGM individuals but not in later hunter-gatherers. At present, the genetic data on Upper Paleolithic, and especially pre-ice age, populations are too sparse to comment on whether or not this is representative of a change in the genetic structure of the population, perhaps caused by a bottleneck during the LGM and a subsequent repopulation from glacial refugia.
Using ancient mtDNA sequences from securely dated archaeological samples as calibration points has allowed us to obtain an estimate of the mtDNA substitution rate that is complementary to the existing estimates based on calibration from the fossil and archaeological records. We arrive at a rate of 1.57 3 1028 substitutions per site per year for the coding region and 2.67 3 1028 substitutions per site per year for the whole molecule, which is approximately 1.6-fold higher than the fossil-calibrated rate. Our inferred substitution rate from the whole mtDNA implies a coalescence date for all modern human mtDNAs of 120–197 kya and of 62–95 kya for hg L3, the lineage from which all non-African mtDNA hgs descend. This places a conservative upper bound of 95 kya for the time of the last major gene exchange between non-African and sub-Saharan African populations. It is important to recognize that this divergence time may merely represent the most recent gene exchanges between the ancestors of non-Africans and the most closely related sub-Saharan Africans and thus may reflect only the most recent population split in a long, drawn-out process of population separation. Nevertheless, the fact that hg L3 is currently so widespread within Africa suggests that the population split dated by L3 is likely to be one of the most important ones in that history of separation, giving rise to lineages that contributed substantial fractions to the ancestry of both present-day sub-Saharan African and present-day non-African populations.
Research-Selection for NovoScriptorium: Maximus E. Niles
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