NEW YORK (GenomeWeb News) – The international research team that sequenced the Neandertal nuclear genome reported today that they have garnered genetic evidence of interbreeding between Neandertals and humans.
In a pair of papers appearing online in Science, the researchers described their efforts to sequence and analyze a draft version of the Neandertal nuclear genome using genetic material isolated from bone fragments from three individuals. The team described preliminary results from the Neandertal draft genome at the American Association for the Advancement of Science annual meeting last year.
Now, their comparison of the Neandertal genomes with the genomes of chimpanzees and contemporary humans from around the world hints at previously unappreciated gene flow from Neandertals to humans outside of Africa: their results suggest one to four percent of non-African human genomes are comprised of Neandertal sequence.
"Neandertals are not totally extinct," senior author Svante Pääbo, director of the Max Planck Institute for Evolutionary Anthropology's evolutionary genetics department, told reporters during a telephone press briefing earlier this week. "In some of us they live on a little bit."
Evidence of Neandertals has been detected in Europe and parts of Asia from about 400,000 years ago up until around 30,000 years ago. But although Neandertals appear to be one of the closest evolutionary relatives to humans, past mitochondrial DNA studies argued against intermixing between Neandertals and humans.
"We came into this as a consortium with a very, very strong bias against gene flow," co-corresponding author David Reich, a population geneticist affiliated with Harvard Medical School and the Broad Institute, told reporters.
Pääbo and his co-workers sifted through 21 different Neandertal bones found in a Croatia, Russia, and Spain, eventually selecting three bones — dated at around 38,300 years old and older — for sequencing.
They then made several libraries from each bone and used the Roche 454 GS FLX/Titanium and the Illumina Genome Analyzer II platforms to sequence suitable Neandertal libraries following quality control steps.
Because most of the DNA in the libraries came from microorganisms, the team used restriction enzymes to preferentially chop up sequences that are over-represented in bacterial DNA.
Overall, they were able to sequence some four billion bases of Neandertal DNA, representing roughly 1.3 times coverage of the genome and some 60 percent of the overall genome sequence. Based on their analyses, the researchers estimated less than one percent of the sequence is contaminated with present-day human DNA.
The team then turned their attention to analyzing the Neandertal genome, developing software to identify and align Neandertal sequence with human and chimpanzee genomes, lead author Richard "Edward" Green explained during the press briefing. Green started working on the project as a post-doctoral researcher at the Max Planck Institute and is now a biomolecular engineering researcher at the University of California at Santa Cruz.
Using this inter-species comparison, the researchers were able to pinpoint where substitutions occurred and estimate sequence divergence between modern humans and Neandertals. Based on their analysis so far, they suggest Neandertals and humans diverged about 270,000 to 440,000 years ago.
By sequencing the genomes of five present-day humans from Southern Africa, West Africa, Papua New Guinea, China, and Western Europe to between four and six times coverage and comparing these with the Neandertal genome, the team found that humans have just a few dozen fixed nucleotide substitutions that affect coding sequences compared with the ancestral sequence found in Neandertals and chimps. And just five genes contained two or more of these changes.
Having the Neandertal genome on hand for comparison also provided insights into regions of the human genome showing evidence of positive selection. Among them: parts of the genome housing genes involved in cognition, energy metabolism, and skeletal development.
"I think those results are interesting in that they really point out what we can do with the Neandertal sequence now," Green said.
Perhaps most unexpectedly, the team also found three independent lines of evidence suggesting modern humans interbred with Neandertals an estimated 50,000 to 80,000 years ago, with Neandertal DNA apparently entering the human population after modern humans left sub-Saharan Africa.
For example, when they compared African and non-African genomes without looking at the Neandertal sequence, the researchers found a dozen of anciently divergent regions. Their subsequent search identified ten of these regions in the Neandertal genome.
"Neandertals are slightly more similar in their genome to people outside of Africa," Pääbo said. "It's certainly an indication of what went on socially when humans and Neandertals met."
So far this Neandertal ancestry does not seem to cluster in any one part of the human genome. Nor is it more common in and around protein-coding genes. Instead, Neandertal sequences seem to be randomly dispersed in the genomes of non-Africans.
"It's a small but very real proportion of ancestry in non-Africans today," Reich said.
But although many of the Neandertal remains found so far come from Europe and western Asia, the team found similar levels of Neandertal DNA in individuals from Europe, China, and Papua New Guinea, suggesting the interbreeding they detected may have occurred early in the migration out of Africa, possibly in the Middle East or North Africa.
The extent of the Neandertal-human interbreeding is unclear, since the same genetic signature could be observed with a small number of events in a small founder population or frequent intermixing in a large population, Reich noted.
In another arm of the project, researchers used targeted hybridization capture to compare around 14,000 protein-coding sequences for Neandertal with 50 modern humans, identifying 88 fixed amino acid substitutions in humans compared with Neandertals.
Pääbo estimates the team has spent roughly €2 million to €3 million ($2.5 million to $3.9 million) sequencing the Neandertal genome, though he noted that sequencing for the next stage of the project should be both cheaper and faster. The researchers ultimately hope to get 10 to 20 times coverage of the Neandertal genome.