Advances in genetic research enhancing our understanding of human evolution

By David Warmflash, MD | 13 May 2016
Genetic Literacy Project

(Credit: Gary Todd / Flickr / Public Domain)

The last few years have brought an increasing number of stories in popular media about extinct hominids, ancient DNA, and human evolution and this parallels expansion of research studies in the scientific literature. In April, for instance, came a story about the European wipe-out of early Americans that depended totally in genetic sequence analysis. In March, there also was a lot of discussion surrounding newly published studies concerning ancestors of modern humans interbreeding with ancient human species that are now extinct.

One study, published in the prestigious journal Science and covered in the New York Times, presented evidence that there were a minimum of four instances of admixing of genes from Neanderthals and other extinct human species into the modern human gene pool. Also getting a lot of attention, a Harvard/UCLA study published in the journal Cell Biology demonstrated that many people, particularly those from southeast Asia, have more sequences from another ancient human species, the Denisovians, than they have from Neanderthals.

The two studies added to a growing awareness of human interspecies mixing tens of thousands of years ago. It’s an idea that has complicated the older view that modern humans, Homo sapiens sapiens, completely replaced other human species by about 30,000 years ago, but the complexity does end with the fact that there was admixing, and that also received attention in the news. This month, for example, there was a big story, based on a Stanford University study, about how, despite interbreeding way back when, modern men lack Y chromosome genes from Neanderthals. This does not mean that Neanderthal men did not start paternal lines that persisted through modern human populations, but if they did then their Y chromosome genes eventually disappeared.

Going back one, two, and three years, there have been story after story about Neanderthals, Denisovians, ancient DNA, and early human species in general. So what’s happening? Are we going through some kind of hominid fad, or are paleoanthropologists actually making discoveries with increasing speed? The details in the science literature suggests that it’s the latter. They are progressing more rapidly than in the past. New data are coming in with increasing frequency and this has to do with advances in molecular genetics, especially in technology that’s being applied to extracting and sequencing of ancient DNA.

An expanding human family

Humanity’s closest living relatives are chimpanzees, but the species Homo neanderthalensis is the most famous example as an extinct but much closer cousin or our own species. H. neanderthalensis has been widely publicized, because scientists have been excavating Neanderthal skulls, skeletons, and tools since the 19th century, and even prior to modern genetic analysis there was enough of a fossil record to reveal something very intriguing—that Neanderthals coexisted with modern humans until about 30,000 years ago. Both also coexisted with a more ancient species called Homo erectus, which migrated out of Africa prior to the emergence of the ancestral line that lead to H. sapiens. H. erectus had a brain size much larger than that of a still more ancient group of hominids called Australopithecines, but much smaller than the brains of modern humans and Neanderthals. The brain size was one of several anatomic features that were intermediate between modern humans and the Australopithecines (which were more like upright apes than like humans), so H. erectus emerged as the candidate common ancestor of H. neanderthalensis, H. sapiens, and additional ancient human species that were discovered in the 20th and 21st centuries.

One of those other ancient human species was Homo heidelbergensis, also called Homo rhodesiensis), bones of which were discovered gradually throughout the 20th century. H. heidelbergensis had features intermediate between H. erectus and H. sapiens, including a brain size almost as big as that of H. sapiens. This suggests that H. heidelbergensis, rather than H. erectus, could represent the more recent common ancestor of modern humans and Neanderthals.

(Credit: Prof saxx / Wikimedia Commons / Public Domain)

Whether modern humans and Neanderthals diverged from H. heidelbergensis or H. erectus is debated and that issue contributes to the stories on human ancestors published over the last couple of decades. Also, fossils of new species were discovered during the first decade of the 21st century, all of which coexisted with modern humans and Neanderthals. One is Homo floresiensis, which had a very small brain and another is Homo sapiens denisova; that’s that Denisovians that have been so hot news cycles lately. Denisovian man is a popular topic for the public, because large segments of the modern human population possess more Denisovian sequences than Neanderthal sequences, but it also represents a scientific, technological milestone based on how it was discovered. First it was a tiny finger bone in Siberia in 2008—the tip of the pinky—and more recently a Denisovian toe bone was found. From that pinky bone, researchers extracted DNA, separated out the DNA that was from soil bacteria and other contaminants, and accessed the DNA that had belonged to the ancient human, about 3 percent of the entire DNA sample. They sequenced it and compared it with sequences of modern humans and Neanderthals. That was possible because, the genetic database of Neanderthals has been growing substantially since 1997, when DNA from Neanderthal bone was first extracted and sequenced successfully. As for the result, sequence comparison showed that the pinky bone belonged to a human of an entirely different species. Thus, H. sapiens denisova became the first human species discovered by way of molecular genetics, rather than by comparative bone anatomy.

Improving technology

Clearly, the discovery of fossils of more than one previously unknown human species during this century is fascinating enough to drive research, science journalism, and public interest. But what’s really driving the studies of ancient humans is the ability to analyze molecular fossils—DNA sequences. The technology has advanced considerably since 1997, when a team led by geneticist Svante Pääbo extracted mitochondrial DNA (mtDNA) from Neanderthal bone. mtDNA is the DNA contained within the cell’s energy organelles called mitochondria, which are inherited completely through maternal lineage with no sequence recombination during mating. That makes mtDNA a very straightforward way to make family trees of different species, but the reason why Pääbo used it in 1997 was that it’s easier to obtain from an ancient bone than nuclear DNA, the DNA that’s inherited from both parents. Unlike the nucleus, which is present in one copy, each bone cell has hundreds of mitochondria, so it has hundreds of copies of the same mitochondrial genome, and that’s why technology for obtaining and using mtDNA was available first.

By comparing variations between the mtDNA of modern humans and Neanderthals, Pääbo and colleagues demonstrated an evolutionary divergence between the two species occurring up to approximately 500,000 years ago. This was with no mixing of mitochondrial genes since the time of that divergence, so when the study came out in 1997 there was not yet any molecular evidence for the interspecies mixing that we’ve been hearing about over the last several years. To obtain such information, researchers would need to recover and sequence nuclear DNA, an achievement that Pääbo in the years following his mtDNA success guessed would be almost impossible—until a few years later when technology actually did make it possible, leading to family trees that were not as crisp as the ones made from mtDNA. In other words, the nuclear DNA comparisons, showed some contribution by Neanderthals to the modern human genome, and today we know that it happened at least four times and that mixing occurred with Denisovians too.

Over the years, the advances leading to mtDNA and nuclear DNA comparisons have depended on new techniques for extracting DNA from the bone. The technology for recognizing and screening out contaminating DNA has been improving as well. That’s vital because if you are lucky enough to find DNA in a Neanderthal bone sample, most likely it will be bacterial DNA, plus there can be contaminants from various animals, including modern humans. Finally, various computational techniques are needed, not just to compared sequence differences, but also to recognize features in ancient DNA that are not the result of evolution and divergence, but merely from chemical reactions that occur over time. These chemical reactions change the nucleotide bases that comprise the letters of the genetic alphabet. Advances in computing power and clever software have helped address those problems, but another major factor that’s come into play in just the last few years is the sequencing technique itself. Today, ancient DNA, can be sequenced much faster than it could be in the 1990s.

Evolution of sequencing

The gold standard method for reading nucleotide base sequences in DNA is called Sanger sequencing, invented by double Nobel Prize winner, biochemist Frederick Sanger in the 1970s. It works using chemical methods that identify which of the four DNA bases—A,C,G, or T—is located at one end of a DNA segment, an enzyme that creates segments of varying length based on the unknown DNA sequence, and a technique called electrophoresis, which separates the newly created segments based on their lengths. Until about 1990, the process was manual, with researchers going through a process of pipetting, heating, cooling, mixing sample, and preparing and running electrophoresis gels over and over. This would give them the sequences of little pieces of the unknown DNA, which then they’d piece together. As you can imagine, it was a long process just to obtain the sequence of one small gene, but this changed with the use of a method called capillary electrophoresis and increasingly high levels of automation.

By the time of landmark 1997 Neanderthal study, Sanger sequencing had advanced to the point that it was routine, reliable, and fairly quick for reading the sequence of mystery DNA, so it was being applied to an increasingly wide range of applications. That could include solving crimes or just about anything in science involving biological samples. While Sanger sequencing requires a certain minimal amount of DNA, another process called polymerase chain reaction (PCR) was developed in the 1980s and 90s that could amplify small amounts of DNA into enough copies for the sequencing to work. That was vital for people who wanted to know the sequence of ancient DNA from a bone, where the yield is almost always going to be extremely low. Even so, it was just barely enough to make mtDNA studies possible, which is why obtaining and sequencing nuclear DNA from an ancient bone enough for genome comparisons seemed like science fiction.

(Credit: Shutterstock)

But over the years, new techniques that vary from the Sanger method have been coming on line. Collectively, they’re known as next generation sequencing (NGS). Unlike the Sanger method, which was manual initially and had automation adapted to it, NGS methods have been created specially for use in automated systems. They differ in certain aspects and some are optimal for specific sequencing equipment produced by specific companies, but what they all have in common is capability for very high throughput. Sanger with capillary electrophoresis is the gold standard for sequencing accuracy and it is still a good choice for various applications—for instance, sequencing a single gene, or the genome of a bacterium. But while NGS has certain technical disadvantages for certain applications, those disadvantages come into play mostly when nice, long, high quality piece of DNA is available. That’s not the case at all with ancient DNA, and so, over the last decade, NGS has been applied increasingly to paleoanthropology. That’s been generating so many data from small amounts of genetic material, whether its mtDNA or even nuclear DNA from a mandible or finger bone, or something else. All of this has led to an increasing number of good, fascinating studies, and given the continuing progression of genetic technology we can expect to learn a whole lot more about ancient humans in the years to come.

Reprinted with permission from the author.

David Warmflash is an astrobiologist and science writer. He received his M.D. from Tel Aviv University Sackler School of Medicine, and has done post doctoral work at Brandeis University, the University of Pennsylvania, and the NASA Johnson Space Center, where he was part of the NASA’s first cohort of astrobiology training fellows. Dr. Warmflash has written numerous articles covering a range of science topics, from the search for extraterrestrial life and space exploration to the origins of life, genetics, neuroscience, biotechnology, and the history of science. His articles have appeared in various publications, including Wired UK, Discover, Scientific American, Genetic Literacy Project, and Cricket Media. Throughout 2018, he did a blog post series on the emergence of ancient science for Vision Learning, covering thinkers from history. Many of these ancient pioneers of science also make an appearance in his book, “Moon: An Illustrated History: From Ancient Myths to the Colonies of Tomorrow”. Follow him on Twitter @CosmicEvolution.

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