Virtually all complex cells – better known as eukaryotes – have at least two separate genomes. The main one sits in the central nucleus. There’s also a smaller one in tiny bean-shaped structures called mitochondria, little batteries that provide the cell with energy. Both sets of genes must work together. Neither functions properly without the other.

Mitochondria came from a free-living bacterium that was engulfed by a larger cell a few billion years ago. The two eventually became one. Their fateful partnership revolutionised life on this planet, giving it a surge of power that allowed it to become complex and big (see here for the full story). But the alliance between mitochondria and their host cells is a delicate one.

Both genomes evolve in very different ways. Mitochondrial genes are only passed down from mother to child, whereas the nuclear genome is a fusion of both mum’s and dad’s genes. This means that mitochondria genes evolve much faster than nuclear ones – around 10 to 30 times faster in animals and up to a hundred thousand times faster in some fungi. These dance partners are naturally drawn to different rhythms.

This is a big and underappreciated problem because the nuclear and mitochondrial genomes cannot afford to clash. In a new paper, Nick Lane, a biochemist at University College London, argues that some of the most fundamental aspects of eukaryotic life are driven by the need to keep these two genomes dancing in time. The pressure to maintain this “mitonuclear match” influences why species stay separate, why we typically have two sexes, how many offspring we produce, and how we age.

Dancing out of step

Here’s the problem: both sets of genes help to create proteins that sit in the mitochondria and carry out one of the most important of chemical reactions: respiration. The proteins strip electrons from our food and pass them along from one to another. They eventually deposit the electrons onto oxygen; this produces water and releases energy. These ‘electron transfer chains’ are the stuff of life, and they only work if the proteins involved are built correctly.

The proteins in the chain are made of different subunits. Some are built using instructions from nuclear genes, while others are built using mitochondrial genes. They different parts must fit together with nanometre precision. Even a small change in their shape will produce botched proteins that fumble their electrons. If fewer electrons make it to the end of the chain, the mitochondria produce less energy. The leaking electrons can also react with oxygen directly to produce destructive molecules called free radicals.

So, cells with mismatched nuclear and mitochondrial genes face a double whammy of less energy and leaking free radicals. This has two important consequences for the evolution of eukaryotes: it creates a barrier between different species, and it favours the evolution of two sexes.

Within a species, the nuclear and mitochondrial genomes have adapted alongside one another so that their protein components seamlessly fit together. These dancers don’t swap their partners easily. If different species mate, they destroy this exquisite co-evolution, which might explain why hybrids encounter so many problems. Respiration is difficult for them, their mitochondria can’t produce any energy, they’re bombarded by leaking free radicals, and many of their cells top themselves. With so many problems, it’s no wonder that many hybrids become sterile or weak, or fail to develop properly at all. This is the price of a mitonuclear mismatch.

Why two sexes?

Mismatches can be easily weeded out by natural selection because every individual has the same mitochondrial genome in all of its cells. Those that match up well with the nuclear genome will survive; those that match poorly will die. This weeding process breaks down if individuals have many different types of mitochondria. In this scenario, the bad matches cancel out the good ones in any individual, and everyone ends up being decidedly average. Natural selection has little to work with.

Over time, individuals with a single mitochondrial genome will do better than those with many. The fittest of them will thrive thanks to natural selection, while their peers stagnate. Lane argues that one of the easiest ways of ensuring that an individual has a uniform set of mitochondria is to have two sexes. One (usually female) hands down an identical set of mitochondria to its young, and the other (usually male) doesn’t. That’s a major difference between the two sexes; some (including Lane) would argue it’s the main difference.

There are species that do things differently, but they are exceptions that prove the rule. Some slime moulds have 13 different sexes, but after mating, they destroy all but one set of mitochondria. Some fungi, like baker’s yeast, inherit mitochondria from both parents, but they are quickly separated so that individual cells only contain one type.

Here’s the gist: mitonuclear mismatches are easier to weed out if individuals test-drive one set of mitochondrial genes against one set of nuclear genes. And having two sexes is an easy way to do that.

The death threshold

The threat of mitonuclear mismatch can also explain the different lifestyles of different species. Mismatches cause a leak of free radicals and cells have two ways of dealing with that. If the leak is fairly minor, the cell can make more mitochondria to compensate. If the leak is severe enough, the cell commits suicide through a process called apoptosis. Lane’s idea is that there’s a threshold that determines which route a cell will take – a level of leakage where it chooses to cut its losses rather than fix the problem.

Different species set their ‘apoptotic threshold’ at different levels. For example, birds and bats need a lot of energy to fly, and their nuclear and mitochondrial genomes must match perfectly. The proteins of their mitochondria have to shunt electrons from one to another quickly and efficiently. Even slight mistakes would compromise their energy levels, and that can’t be tolerated.

So, birds and bats have very low leak thresholds. Even a slight trickle of free radicals betrays the fact that their two genomes aren’t meshing properly – time for their cells to die. Dying cells mean dying embryos, and many are eliminated before they fully develop. Only a precious few would make it through this harsh selection process. Lane thinks that this could explain why these species tend to have low fertility rates and few offspring.

By contrast, a rat has less demanding energy needs, and the electron transfer chain in its mitochondria can afford to be leakier and less efficient than that of a bird. The rat can handle a poorer mitonuclear match, so it sacrifices fewer embryos on the altar of perfection. It follows that rats are also more fertile, and produce larger litters.

Ageing apart

Even well-matched nuclear and mitochondrial genomes don’t stay that way forever. As individuals age, leaking radicals will damage and mutate the mitochondrial genome, ruining its match with the nuclear one, and causing even heavier leaks. This happens, even if the initial stream of radicals is small. As time wears on, the dancers inevitably fall out of step with each other. You can see this if you compare young and old tissues: the young cells will all have genetically identical mitochondria, while those in the old cells will be a mix of different mutants.

As more cells pass the tolerance threshold, more of them die. Tissues that use the most energy, like the muscles and brain, have the heaviest leaks and wear away faster. Meanwhile, the surviving cells experience even greater energy demands. They enter a downward spiral with sweeping consequences: they leak free radicals like sieves; their DNA becomes more fragile; their genes become switched on in different ways; they release chemicals that trigger inflammation. In short, they create the perfect set-up for cancer, heart disease, diabetes, Alzheimer’s and many of the other diseases of old age.

Almost all of the major traits of ageing can be predicted by a growing rift between two genomes, and a widening leak of free radicals. The leak worsens with time, so tissues die, especially gas-guzzling ones. Those that survive are more likely to become diseased. And the fast the leak, the faster all of this happens. This explains why species that tolerate less free radical leaks tend to enjoy longer lives. Consider pigeons and rats: both species are similar in both size and metabolic rates, but pigeons have far lower rates of leaking electrons in their mitochondria. They also live ten times longer.

A simple idea

For now, this is all a grand hypothesis, albeit one that is grounded on a lot of existing evidence. Lane now wants to explore ways of testing his idea. The most obvious first step would be to see if there actually is a lea threshold that varies between cells. It should be straightforward to measure the extent of free radical leaks in cells, and the level that makes them kill themselves.

He also wants to look at species with high energy needs like birds, to see whether a large proportion of their embryos are being lost. He’s also interested in how this applies to humans. “It would be interesting to get data from fertility clinics to see if there are any groups or populations that struggle to conceive,” he says, “and if any of this can be put down to incompatibilities between mitochondria and nuclear backgrounds. Around 40% of pregnancies end in miscarriage and we don’t know why.”

There is compelling majesty to Lane’s idea. At its heart, it is deceptively simple: we have two genomes that need to work together, and you can tell how well they’re doing this by the strength of the free radical leak. From that simple concept, you can logically derive how fitness, fertility and lifespan are linked in different species. You can also predict the process of ageing and the onset of age-related diseases within individuals.

“A lot of this has to be true on logical grounds,” says Lane. “We know that there is co-adaptation between these two genomes and many predictions emerge seamlessly from some simple reflections on that process. The big question is whether it’s important in the greater scheme of things.”

Reference: Lane, N. (2011). Mitonuclear match: Optimizing fitness and fertility over generations drives ageing within generations BioEssays DOI: 10.1002/bies.201100051

Lane, N. (2011). The Costs of Breathing Science, 334 (6053), 184-185 DOI: 10.1126/science.1214012

Image: by Ticipico

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It couldn’t be easier to make sweeping edits on a computer document. If I were so inclined, I could find every instance of the word “genome” in this article and replace it with the word “cake”. Now, a team of scientists from Harvard Medical School and MIT have found a way to do similar trick with DNA. Geneticists have long been able to edit individual genes, but this group has developed a way of rewriting DNA en masse, turning the entire genome of a bacterium into an “editable and evolvable template”.

Their success was possible because the same genetic code underlies all life. The code is written in the four letters (nucleotides) that chain together to form DNA: A, C, G and T. Every set of three letters (or ‘codon’) corresponds to a different amino acid, the building blocks of proteins. For example, GCA codes for alanine; TGT means cysteine. The chain of letters is translated into a chain of amino acids until you get to a ‘stop codon’. These special triplets act as full stops that indicate when a protein is finished.

This code is virtually the same in every gene on the planet. In every human, tree and bacterium, the same codons correspond to the same amino acids, with only minor variations. The code also includes a lot of redundancy. Four DNA letters can be arranged into 64 possible triplets, which are assigned to only 20 amino acids and one stop codon. So for example, GCT, GCA, GCC and GCG all code for alanine. And these surplus codons provide enough wiggle room for geneticists to play around with.

Farren Isaacs, Peter Carr and Harris Wang have started to replace every instance of TAG with TAA in the genome of the common gut bacterium Escherichia coli. Both are stop codons, so there’s no noticeable difference to the bacterium – it’s like replacing every word in a document with a synonym. But to the team, the genome-wide swap will eventually free up one of the 64 triplets in the genetic code. And that opens up many possible applications.

“We are actively pursuing three of them,” says Isaacs. First, they could assign the empty triplet to unnatural amino acids that sit outside the standard twenty. “This [could] expand the diversity of possible enzymes and create new classes of drugs, industrial enzymes and biomaterials.”

Second, the team could use the tweaked genetic codes to make living things resistant to viruses. Viruses make copies of themselves by hijacking the protein-making factories of their hosts. They depend on the fact that their proteins are encoded by the same triplets as those of their hosts. If their hosts stray from this universal genetic code, their factories will mangle the virus’s instructions, creating distorted and useless proteins. That would be useful for industry as well as medicine. The biotechnology company Genzyme had to shut down a manufacturing plant for several months after it was hit by a contaminating virus. Millions of dollars were lost.

Third, and for similar reasons, the altered codes could be used to contain genetically modified organisms, preventing them from breeding with wild populations. It’s the geneticist’s version of the Tower of Babel story – modified creatures would be imprisoned by their own genetic tweaks, unable to productively exchange genes with natural counterparts.

All three applications are some distance away in the future, but Isaacs, Carr and Wang have taken an important step towards them. Their genome-wide edits relied on two complementary technologies, invented by their team – MAGE, which substitutes TAA for TAG in separate pieces of bacterial DNA, and CAGE, which knits the pieces together into a whole genome.

MAGE, the older of the two techniques, made its debut two years ago. It stands for “multiplex automated genome engineering”, a fancy way of saying that it can easily change a genome many times over. It was originally used to create millions of small variants of bacterial genomes, producing a multitude of strains that can be tested for new abilities. As Jo Marchant puts it in her excellent feature, it’s an “evolution machine”. In its debut, within a matter of days, it had evolved a strain of E.coli that would produce large amounts of lycopene, a pigment that makes tomatoes red.

MAGE is a versatile editor. Not only can it create many diverse changes in a group of cells, it can also create many specific changes in a single cell. That’s what Isaacs, Carr and Wang have now done. TAG appears in 314 places throughout the E.coli genome as a stop codon. For each one, the team created a small stretch of DNA that had TAA instead of TAG, surrounded by exactly the same letters. They fed these edited fragments into bacteria, which used them to build new copies of their own DNA. The result: daughter bacteria with edited genomes.

In this way, Isaacs, Carr and Wang created 32 strains of E.coli that, between them, had every possible substitution of TAG to TAA. This might seem overly complicated, but replacing every TAG with TAA in a single step would be inefficient, slow, and error-prone. A single mistake could be lethal for the microbes. By taking things slowly, and spreading the substitutions among 32 strains, the team could better troubleshoot any tricky snags.

To combine the 32 strains into one, Isaacs, Carr and Wang developed CAGE (or “conjugative assembly genome engineering”). The technique relies on the bacterial equivalent of sex – a process called conjugation where two cells sidle up, form a physical link between one another, and swap DNA.

The team matched their 32 strains up in pairs, in a league that looked like a knock-out sports tournament. One strain of each pair would deliver its edited genes into its partner, and the incoming genes were designed to merge with those of the recipient in specific ways. Thirty-two strains with 10 edits each became sixteen strains with 20 edits each. Sixteen turned into eight and eight into four.

At the time of publication, the team had reached this “semi-final” stage. They had four strains of E.coli, each with a quarter of its genome stripped of TAG codons. The strains seem to be growing normally, proving that, individually at least, the TAG codons aren’t necessary for the bacterium’s survival. Whether E.coli can survive without any TAG codons at all is still unclear, but the team suspects this will be the case. If so, they’ll set about reprogramming the unused TAG codon to represent an unusual amino acid beyond the normal set of 20.

Why publish a paper at the semi-finals? “It is indeed an odd stopping point,” admits Carr. “[We’ve] been working on this project for 7 years and we decided to publish at this point largely because we have so much to talk about: the successful innovation of the CAGE technology and it’s integration with MAGE for genome engineering at large and small DNA scales. If you dig into the supplemental data of this paper, there’s another 1-2 more papers worth of stuff in there.

Isaacs points out that only one other research group is “working on genome engineering at this scale”: the J. Craig Venter Institute (JCVI). Last year, they made headlines by creating a bacterial genome, 1.1 million DNA letters (base pairs) long, and implanting it into the shell of a different bacterium.

Isaacs says, “[They] took 10 articles to get to a slightly-modified one million base pairs. We hope to get to a highly-modified, industrially useful 4.7 million base pair genome in three papers.” That includes the one that introduced MAGE to the world in 2009, and the current one that couples it with CAGE. The third one, due in the next year or so, will complete the trilogy – it will feature the final strain, . “All the pieces are in place,” says Carr. “We have a high degree of confidence we will reach our goal.”

What does the JCVI make of this? In a statement released to the press, Dan Gibson and Craig Venter point out that the MAGE/CAGE method still requires an existing genome to work from. Replacing an entire codon is a remarkable achievement, but it’s still a tweaking game. The end result will still be a genome that’s at least 90% similar to the original one. Gibson and Venter say, “Ultimately, we at JCVI would like to design cells from scratch.” The only way to do this is to synthesise an entirely fresh genome, rather than modify an existing one.

They add, “We continue to believe there will be and must be many different techniques developed to engineer and construct genomes so that the field can mature, allowing new and important products to be made. We believe the Isaacs et al paper is a positive addition to the field.”

Reference: Isaacs, Carr, Wang, Lajoie, Sterling, Kraal, Tolonen, Gianoulis, Goodman, Reppas, Emig, Bang, Hwang, Jewett, Jacobson & Church. 2011. Precise Manipulation of Chromosomes in Vivo Enables Genome-Wide Codon Replacement. http://dx.doi.org/10.1126/science.1205822

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In September 1941, Hu Chengzhi placed several skulls into two wooden crates. Around him, China was at war with Japan, so he was sending the skulls to the US for safekeeping. They never arrived. Hu was among the last people to see one of the most important palaeontological finds in history. These lost skulls belonged to Homo erectus pekinensis, known as Peking Man.

You can read all of them free online, which include the Maxberg Archaeopteryx, Nixon’s moon rocks, the recipe for Damascus steel and moon trees.