Hares And Tortoises

You may have noticed that the last few months have seen a bit of a DNA-fest in these pages. Don’t blame me. It’s all the fault of them scientists beavering away in their labs. We’ve just done “Making Movies in DNA“, in “And Now There Are Six!!” the genetic code was expanded from four to six units by making two new ones artificially and in “How Does DNA Do It?” we saw how words can be transformed into a sequence of DNA.

Now they’re at it again – or at least Stephen Kowalczykowski, James Graham and colleagues of the University of California at Davis are – revealing yet more astonishing things about this molecule, just when you could be thinking we’ve got the hang of it.

I might add that I’m grateful to my correspondent David Archer of The Society of Biology for bringing this piece of work to my notice as I’d missed it in the journal Cell (cries of ‘shame’ and ‘shurely shome mistake’ mingle in the background).

What is it this time?  

Well it’s two really astonishing things about DNA replication – the process by which double-stranded DNA is pulled apart so that each strand can act as a template for making a new DNA molecule. Result: as cells progress towards division, they double their DNA content so that equal amounts can be given to each new daughter cell. The first source of amazement is that Stephen K & chums have filmed this happening in real time. That’s a terrific feat – but what it reveals is quite bizarre.

Up to now it’s been assumed that the protein machines (DNA polymerases) doing the biz trundle along each of the separated strands of parental DNA at more or less the same speed. It would seem to make no sense to do otherwise and risk ending up with the job half done. In other words, the duplication of the two strands is coordinated. Is that what K & Co found? Not a bit of it! Extraordinary to relate, it appears that there’s no coordination between the strands at all!! Not for the first time in the history of molecular biology a technical advance has thrown up the totally unexpected. Before we look at the results in a bit more detail, a little background might be useful.

One divides into two

Making two identical copies of DNA from one original happens every time one cell divides to make two. And there’s a lot of it about. As is well known, we all start out as one cell (i.e. a fertilized egg) that turns into a human being – 50 trillion cells (that’s 5 + 13 zeroes). And even after we’ve been assembled it takes a lot of cell-making to keep us ticking over – about one million new cells every second. Just take a second to think about that: DNA comes in the well-known form of a double helix – two strands made up of chemical units (called nucleotides) linked together. Each unit has one of four bases (cytosine (C), guanine (G), adenine (A), or thymine (T)) and the strands are “complementary” because C pairs with G and A with T – a rigid rule that means if you know the sequence of bases in one strand you can work out what it is in the other. So far so simple. But, as we noted in “How Does DNA Do It?”, the coding power of DNA lies in its size. In us three billion letters are available to do the encoding. That is, there are just over 3,000 million units in each chain – i.e. 3,000 million base-pairs all told. And all of these are copied (twice) for every new cell.

DNA replication: The double helix is ‘unzipped’ so that each separated strand (turquoise) can act as a template for replicating a new partner strand (green). This creates a ‘replication fork’ – two branches of single stranded DNA. The new strands are made by protein complexes called DNA polymerases chugging along the parent strands, making new, complementary, strands as they go. There’s a small technical wrinkle here: new DNA chains can only be extended in one direction. This means that, while one strand can be made continuously (the leading strand), the other has to be put together in short bits as the parent strand is unwound, with the bits being joined up afterwards (the lagging strand).

 

 

Timing is everything

So the cell’s task is to unzip the double helix and use each exposed strand as a template for building a new partner strand. Things are helped by DNA being split into fragments (chromosomes: 23 pairs in humans + 2 sex chromosomes, 46 per cell all told). Even so, chromosomes are huge: the longest (chromosome 1) has nearly 250 million base-pairs; the shortest (chr 21) has about 47 million. The problem for the machinery that has evolved for the job is that it cranks along at 50 pairs per second – roughly a month per chromosome. But in a normal cell cycle the whole business is done in about two hours! That’s made possible because replication doesn’t do the obvious: start at one end and work its way to the other. Cunningly it hits lots of ‘start points’ – up to 100,000 in a single cell – making lots of short bits at the same time that are then joined up. In other words replication proceeds simultaneously from many different sites in chromosomes. Enzymes join the pieces together to make the final, complete copy.

It’s rather like you having some horribly repetitive chore to do – washing up after a big dinner. On your own you might start at one end of the pile and work through it but, far better, get one member of the family to do the plates, another the cutlery, etc. and – job done!!

Now for today’s bit of amazing science

What Kowalczykowski and friends did was to extract DNA from bugs (E.coli bacteria in fact, that can make DNA about 20 times faster than human cells), set up a replication system and measure what went on by microscopy, using a dye (SYTOX Orange, which is fluorescent) that sticks to complete double helices but not to single strands. Thus they could track progress along a strand as a new double helix formed. What they saw was that each strand acted independently of the other. Overall, the rate of replication of the two strands was about the same (as it must be in the end) but along the way there were stops and starts and sometimes one strand would grow at ten times the speed of the other. How weird is that?!!

Seeing DNA being made. In this picture microscopy reveals three extending stretches of double-stranded DNA being made (Graham et al. 2017). Click here to see video.

You could picture DNA replication as one of those Swiss railway trains cranking up a mountain at an improbable angle, using a rack-and-pinion to stop it sliding backwards. Think of the engaging cogs as new base-pairs. The train just keeps chugging along until it reaches the its next stop. But why doesn’t the DNA-making machinery do the same? Well, we haven’t much of a clue. One difference is that the train has its track (and rack) laid out before it, whereas DNA is continuously being unwound to open the template. Some bits are more difficult to unwind than others and this variation may cause the system to go in fits and starts. Another contribution many come from the many proteins involved in this complicated process. As well as the polymerases there are things that unwind DNA, stabilize it, stitch new bits together, etc. and these complexes are continuously forming, falling apart and re-assembling – all of which gives plenty of scope for erratic behaviour.

Fact of the matter is, we don’t know. So, in revealing completely unexpected behaviour, this technical triumph throws up the question of how two strands working independently manage, in the end, to come up with the perfect finished product.

But hey! This wouldn’t be science if we had all the answers!

Reference

Graham, J.E., Marians, K.J., Kowalczykowski, S.C. (2017). Independent and Stochastic Action of DNA Polymerases in the Replisome. Cell 169, 1201–1213.

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Making Movies in DNA

Last time we reminded ourselves of one of the ways in which cancer is odd but, of course, underpinning not just cancers but all the peculiarities of life is DNA. The enduring wonder is how something so basically simple – just four slightly different chemical groups (OK, they are bases!) – can form the genetic material (the instruction book, if you like) for all life on earth. The answer, as almost everyone knows these days, is that there’s an awful lot of it in every cell – meaning that the four bases (A, C, G & T) have an essentially infinite coding capacity.

That doesn’t make it any the less wonderful but it does carry a huge implication: if something you can squeeze into a single cell can carry limitless information it must be the most powerful of all storage systems.

A picture’s worth a thousand words

We looked at the storage power of DNA a few months ago (in “How Does DNA Do It?”) and noted that its storage density is 1000 times that of flash memories, that it’s fairly easy to scan text and transform the pixels into genetic code and that, as an example, someone has already put Shakespeare’s sonnets into DNA form.

Now Seth Shipman, George Church and colleagues at Harvard have taken the field several steps forward by capturing black and white images and a short movie in DNA. Moreover they’ve managed to get these ‘DNA recordings’ taken up by living cells from which they could subsequently recover the images.

Crumbs! How did they do it?

First they used essentially the text method to encode images of a human hand: assign the four bases (A, C, G & T) to four pixel colours (this gives a grayscale image: colours can be acquired by using groups of bases for each pixel). These DNA sequences were then introduced into bacteria (specifically E. coli) by electroporation (an electrical pulse briefly opens pores in the cell membrane).

The cells treat this foreign DNA as though it was from an invading virus and switch on their CRISPR system (summarized in “Re-writing the Manual of Life”). This takes short pieces of viral DNA and inserts them into the cell’s own genome in the form of ‘spacers’ (the point being that the stored sequences confer ‘adaptive immunity’: the cell has an immunological memory so it is primed to respond effectively if it’s infected again by that viral pathogen).

In this case, however, the cells have been fooled: the ‘spacers’ generated carry encoded pictures, rather than viral signatures.

Because spacers are short it’s obvious that you’ll need lots of them to carry the information in a photo. To keep track when it comes to reassembling the picture, each DNA fragment was tagged with a barcode (and fortunately we explained cellular barcoding in “A Word From The Nerds”).

Once incorporated in the bugs the information was maintained over many bacterial generations (48 in fact) and is recoverable by high-throughput sequencing and reconstruction of the patterns using the barcodes.

And the movie bit?

Simple. In principle they used the same methods to encode sequential frames.

Pictures in DNA.

Top: Using triplets of bases to encode 21 pixel colours. Images of a human hand (top) and a horse (bottom) were captured. For the movie they used freeze frames taken in 1872 by the English photographer Eadweard Muybridge. These showed that, for a fraction of a second, a galloping horse lifts all four hooves off the ground. Seemingly this won a return for the sometime California governor, Leland Stanford (he of university-founding fame) who had put a wager on geegees doing just that. From Shipman et al., 2017. You can see the movie here.

Getting the picture clear

To recap, in case you’re wondering if this is some scientific April Fools’ prank. What Church & Co. did is scan pictures and transform pixel density into the genetic code (i.e. sequences of the four bases A, C, G & T). They then made DNA carrying these sequences, persuaded bacteria to take up the DNA and incorporate it into their own genomes and, after growing many generations of the bugs, extracted their DNA, sequenced it and reconstructed the original images. By scanning sequential frames this can be extended to movies.

It’s not science fiction – but it is pretty amazing. With a droll turn of phrase Seth Shipman said “We want to turn cells into historians” and the work does have significant implications in showing something of the scope of biological memory systems.

Won’t be long before the trendy, instead of birthday presents of electronic family photo albums, are giving small tubes of bugs!

References

Shipman, S.L., Nivala, J., Macklis, J.D. & Church, G.M. (2017). CRISPR–Cas encoding of a digital movie into the genomes of a population of living bacteria. Nature 547, 345–349.

Flipping The Switch

If you spend even a little time thinking about cancer you’ll have realised that it’s very odd – and one oddity in particular may have struck you. A general rule is that it can arise anywhere in the body: breast, bowel and lung are commonly affected, but the more than 200 different types of cancer pop up in lots of other organs (e.g. brain, pancreas), albeit less often. But what about those places of which you hear almost nothing? For example, it’s very unusual to hear of heart or muscle cancers. Which raises the obvious question of why? Is there something going on in these tissues that counters cancer development – acts in some way to slow down tumour formation? And if there is, shouldn’t we find out about it?

Zuzana Keckesova, Robert Weinberg and their colleagues from the Whitehead Institute for Biomedical Research, Massachusetts Institute of Technology and other centres have been scratching their heads over this for a while and they’ve recently published an answer – or, at least, one of the answers.

Getting energy from food

To see how their result fits into the jigsaw puzzle we need a quick recap on the chemical processes that go on in cells to keep them alive, aka, metabolism. Occurring in almost all organisms, glycolysis is a central metabolic pathway in which a series of chemical reactions breaks down sugars into smaller compounds, the energy released being captured as ATP (adenosine triphosphate). Needless to say, it’s complicated – there’s 10 steps and it took the best part of 100 years to work them out completely.

Prising open the black box

The story began with the French obsession with wine (which by now they’ve shared with the rest of the world, bless ’em), specifically why sometimes wine tastes horrible. So they put Louis Pasteur on the case and in 1857 he showed that it was all to do with oxygen: if air (oxygen) is present during the fermentation process the yeast cells will grow but fermentation (i.e. alcohol production) will decrease. This showed that living microorganisms were needed for fermentation and led Eduard Buchner to extract the enzymes from yeast and show that they were sufficient to convert glucose to ethanol (alcohol). In other words, you could do it all in a test tube.

The cartoon shows sugar crossing a cell membrane (a bilayer of phospholipids). The 10 steps of the glycolytic pathway (red dots) convert glucose to pyruvate that can become lactic acid or cross the membrane (another lipid bilayer) of mitochondria. In these ‘cells within cells’ oxygen is consumed to make ATP from pyruvate. Glycolysis yields 2 ATPs from each glucose. In mitochondria ‘aerobic respiration’ produces 38 ATPs per glucose – which is why they have been called the “powerhouse of the cell”. In yeast, fermentation produces alcohol from pyruvate.

This was a stunning achievement because it showed for the first time that living systems weren’t inaccessible black boxes. You could take them to bits, find out what the bits were and reassemble them into something that worked – and that’s really a definition of the science of biochemistry. The upshot was that by the 1930s through the efforts of many gifted scientists, notably Otto Meyerhof and Gustav Embden, we had a step-by-step outline of the pathway now known as glycolysis.

Enter Otto Warburg

But by this point a chap called Otto Warburg had noticed that something odd happened to metabolism in cancer. He showed that tumour cells get most of their energy from glucose using the glycolytic pathway, despite the fact that it is much less efficient than aerobic respiration (2 to 38 ATPs per glucose). And they do this even when lots of oxygen is available. Which seems like molecular madness.

Warburg was part of an amazing scientific galaxy in the period from 1901 to 1940 when one out of every three Nobel Prize winners in medicine and the natural sciences was Austrian or German. Born in Freiburg, he completed a PhD in chemistry at Berlin and then qualified in medicine at the University of Heidelberg. Fighting with the Prussian Horse Guards in the First World War, he won an Iron Cross and followed that up with the 1931 Nobel Prize in Physiology or Medicine for showing that aerobic respiration, that is, oxygen consumption, involves proteins that contain iron. However, he made so many contributions to biochemistry that he was actually nominated three times for the prize.

His discovery about tumour cells led Warburg to suggest, reasonably but wrongly, that faulty mitochondria cause cancers – whereas we now know that it’s the other way around: metabolic perturbation is just one of the consequences of tumour development.

But if upsetting mitochondria gives tumours a helping hand, how about looking for factors that help to keep them normal – i.e. using oxidative phosphorylation. And the obvious place to look is in cells that don’t multiply – i.e. appear cancer-resistant.

Which is the idea that led Keckesova & Co to a ‘eureka’ moment. Searching in muscle cells from humans and mice they discovered a protein, LACTB, lurking in their mitochondria. When they artificially made LACTB in a variety of tumour cells both in vitro and in mice it inhibited their growth. In other words, LACTB appears to be a new ‘tumour suppressor’.

What does it do?

It turns out that LACTB works in a quite subtle way. It’s only found in mitochondria, not in the main body of the cell, and it plays a part in making the membrane that forms the boundary of the “powerhouse of the cell”. Membranes are made of two layers of phospholipids arranged with their fatty tails facing inwards. They work as regulatable barriers via proteins associated with the membrane that control the passage of small molecules – so, for example, pyruvate that we mentioned earlier uses specific proteins to cross the mitochondrial membrane.

But aside from their attached proteins, the lipids themselves are a complex lot: they have a variety of fatty acid tails and different chemical groups decorate the phosphate heads. This gemisch arises in part because the lipids themselves control the proteins that they surround. In other words, if the lipid make-up of a membrane changes so too will the efficiency of embedded transport proteins. LACTB controls the level of one type phospholipid (phosphatidylethanolamine, PE): when LACTB is knocked out more PE is made. Thus this tumour suppressor affects mitochondrial lipid metabolism and hence the make-up of the membrane, and its normal role helps in blocking tumour development.

Layers of lipids with their tails pointing inwards make up cell membranes (left): proteins (red & blue blobs) control what can cross the membrane. Phospholipids themselves are a complex mixture with a variety of head groups and fatty acid tails (right).

And the method behind the madness?

So in this newly-discovered tumour suppressor we have a way in which mitochondria can be subverted to promote tumours by changing the properties of their membrane. But what’s the point? Why might it be more profitable for cancer cells to get most of their energy via a high rate of glycolysis rather than by the much more efficient route of oxidising pyruvate in mitochondria – a switch often called The Warburg effect.

There seem to be two main reasons. One is that pathways branch off from glycolysis that provide components to make new DNA – greater flow though glycolysis makes those pathways more active too – a good thing if cells are going to reproduce. The second is that making abnormal amounts of lactic acid actually helps tumour cells to survive and proliferate, it stimulates the growth of new blood vessels to feed the tumour and it can make the immune response – the  defence normally mounted by the host against tumours – less effective.

By affecting mitochondrial function, mutations that knock out LACTB can give the Warburg effect a helping hand and – if the great man’s still following the literature – he may have noted with some glee that this finding, at least, is consistent with his idea that it all starts in mitochondria!

Reference

Keckesova, Z. et al. (2017). LACTB is a tumour suppressor that modulates lipid metabolism and cell state. Nature doi:10.1038/nature21408

Cancer GPS?

The thing that pretty well everyone knows about cancers is that most are furtive little blighters. They kill one in three of us but usually we don’t they’re there until they are big enough to make something go wrong in the body or to show up in our seriously inadequate screening methods. In that sense they resemble heart problems of one sort or another, where often the first indication of trouble is unexpectedly finding yourself lying on the floor.

Meanwhile, out on the highways and byways you are about 75 times less likely to be killed in an accident than you are to succumb to either cancers or circulation failure. Which is a way of saying that in the UK about 2000 of us perish on the roads each year. That it’s ‘only’ 2000 is presumably because here your assailant is anything but furtive. All you’ve got to do is side-step the juggernaut and you’ll probably live to be – well, old enough to get cancer.

Did you know, by the way, that ‘juggernaut’ is said to come from the chariots of the Jagannath Temple in Puri on the east coast of India. These are vast contraptions used to carry representations of Hindu gods on annual festival days that look as though walking pace would be too much for them. So, replace the monsters on our roads with real juggernauts! Problem largely solved!!

Flagging cancer

But to get back to cancer or, more precisely, the difficulty of seeing it. After centuries of failing to make any inroads, recent dramatic advances give hope that all is about to change. These rely on the fact that tissues shed cells – and with them DNA – into the circulation. Tumours do this too – so in effect they are scattering clues to their existence into blood. By using short stretches of artificial DNA as bait, it’s possible to fish out tumour cell DNA from a few drops of blood. That’s a pretty neat trick in itself, given we’re talking about fewer than 100 tumour cells in a sea of several billion other cells in every cubic millimeter of blood.

There are two big attractions in this ‘microfluidics’ approach. First it’s almost ‘non-invasive’ in needing only a small blood sample and, second, it is possible that indicators may be picked up long before a tumour would otherwise show up. In effect it’s taking a biochemical magnifying glass to our body to ask if there’s anything there that wouldn’t normally be present. Detect a marker and you know there’s a tumour somewhere in the body, and if the marker changes in concentration in response to a treatment, you have a monitor for how well that treatment is doing. So far, so good.

And the problem?

These ‘liquid biopsy’ methods that use just a teaspoonful of blood have been under development for several years but there has been one big cloud hanging over them. They appear to be exquisitely sensitive in detecting the presence of a cancer – by sequencing the DNA picked up – but they have not been able to pinpoint the tissue of origin. Until now.

Step forward epigenetics

Shuli Kang and colleagues at the University of California at Los Angeles and the University of Southern California have broken this impasse by turning to epigenetics. We noted in Twenty More Winks that an epigenetic modification is any change in DNA, other than in the sequence of bases (i.e. mutation), that affects how an organism develops or functions. They’re brought about by tacking small chemical groups (commonly methyl (CH3) groups) either on to some of the bases in DNA itself or on to the proteins (histones) that act like cotton reels around which DNA wraps itself. The upshot is small changes in the structure of DNA that affect gene expression. You can think of DNA methylation as a series of flags dotted along the DNA strand, decorating it in a seemingly random pattern. It isn’t random, of course, and the target for methylation is a cytosine nucleotide (C) followed by a guanine (G) in the linear DNA sequence – called a CpG site because G and C are separated by one phosphate (p). Phosphate links nucleosides together in the backbone of DNA.

Cancer cells often display abnormal DNA methylation patterns – excess methylation (hypermethylation) in some regions, reduced methylation in others – that contributes to their peculiar behavior. It’s possible to determine the methylation profile of a DNA sample (by a method called bisulfite sequencing).

Kang & Co. developed a computer program to analyse methylation profiles from solid tumours and healthy samples in public databases and compare them to patient DNA of unknown tissue origin.

The peaks represent CpG clusters that characterize normal cells (top) and a variety of cancers. The key point is that the different patterns identify the tissue of origin (from Kang, S. et al., 2017).

The program’s called CancerLocator and in this initial study it was used to test samples from patients with lung, liver or breast cancer. In the modest words of the authors, CancerLocator ‘vastly outperforms’ previous methods – mind you, they struggle to even to distinguish most cancer samples from non-cancer samples. Nevertheless, CancerLocator’s a big step forward, not least because it can detect early stage cancers with 80% accuracy.

It’s also reasonable to expect major improvements as methylation sequencing becomes more extensive and higher resolution reveals more subtle signatures. What’s more, in principle, it should be able to detect all types of cancers – meaning that, after all so many centuries we may at last have a way of side-stepping the juggernaut.

References

Kang, S. et al. (2017). CancerLocator: non-invasive cancer diagnosis and tissue-of-origin prediction using methylation profiles of cell-free DNA. Genome Biology DOI 10.1186/s13059-017-1191-5.

And Now There Are Six!!

Scientists eh! What a drag they can be! Forever coming up with new things that the rest of us have to wrap our minds around (or at least feel we should try).

Readers of these pages will know I’m periodically apt to wax rhapsodic about ‘the secret of life’ – the fact that all living things arise from just four different chemical units, A, C, G and T. Well, from now on it seems I’ll need to watch my words – or at least my letters – though maybe for a while I can leave it on the back burner in the “things that have been but not yet” category, to use the melodic prose of Christopher Fry.

Who dunnit?

The problem is down to Floyd Romesberg and his team at the Scripps Research Institute in California.

Building on a lot of earlier work, they’ve made synthetic units that stick together to form pairs – just like A-T and C-G do in double-stranded DNA. But, as these novel chemicals (X & Y) are made in the lab, the bond they form is an unnatural base pair.

Left: Two intertwined strands of DNA are held together in part by hydrogen bonds. Right top: Two such bonds (dotted lines) link adenine (A) to thymine (T); three form between guanine (G) and cytosine (C). These bases attach to sugar units (ribose) and phosphate groups (P) to form DNA chains. Right bottom: Synthetic X and Y units can also stick together and, via ribose and phosphate, become part of DNA.

After much fiddling Romesberg’s group derived E. coli microbes that would take up X and Y when they were fed to the cells as part of their normal growth medium. The cells treat X and Y like the units they make themselves (A, C, G & T) and insert them in new DNA – so a stretch of genetic code may then read: A-C-G-T-X-T-A-C-Y-A-T-… And, once part of DNA, the novel units are passed on to the next generation.

Science fiction?
If this has you thinking creation and exploitation of entirely new life forms?!!’ you’re not alone. Seemingly Romesberg is frequently asked if he’s setting up Jurassic Park but, as he points out, the modified bugs he’s created survive only as long as they’re fed X and Y so if they ‘escape’ (being bugs this would probably be down the drain rather than over a fence), they die. Cunning eh?!!

Is this coming to a gene near you?
No. It is, however, clear that more synthetic bases will be made, expanding the power of the genetic code yet further. What isn’t yet known is what the cells will make of all this. In other words, the whole point of tinkering with DNA is to modify the code to make novel proteins. In the first instance the hope is that these might be useful in disease treatment. Rather longer-term is the notion that new organisms might emerge with specific functions – e.g., bugs that break down plastic waste materials.

At the moment all this is speculation. But what is now fact is amazing enough. After 4,000 million years since the first life-forms emerged, more than five billion different species have appeared (and mostly disappeared) on earth – all based on a genetic code of just four letters.

Now, in a small lab in southern California, Mother Nature has been given an upgrade. It’s going to be fascinating to see what she does with it!

Reference

Zhang, Y. et al. (2017). Proceedings of the National Academy of Sciences 114, 1317-1322.

Through the Smokescreen

For many years I was lucky enough to teach in a cancer biology course for third year natural science and medical students. Quite a few of those guys would already be eyeing up research careers and, within just a few months, some might be working on the very topics that came up in lectures. Nothing went down better, therefore, than talking about a nifty new method that had given easy-to-grasp results clearly of direct relevance to cancer.

Three cheers then for Mikhail Denissenko and friends who in 1996 published the first absolutely unequivocal evidence that a chemical in cigarette smoke could directly damage a bit of DNA that provides a major protection against cancer. The compound bound directly to several guanines in the DNA sequence that encodes P53 – the protein often called ‘the guardian of the genome’ – causing mutations. A pity poor old Fritz Lickint wasn’t around for a celebratory drink – it was he, back in the 1930s, that first spotted the link between smoking and lung cancer.

This was absolutely brilliant for showing how proteins switched on genes – and how that switch could be perturbed by mutations – because, just a couple of years earlier, Yunje Cho’s group at the Memorial Sloan-Kettering Cancer Center in New York had made crystals of P53 stuck to DNA and used X-rays to reveal the structure. This showed that six sites (amino acids) in the centre of the P53 protein poked like fingers into the groove of double-stranded DNA.

x-ray-picCentral core of P53 (grey ribbon) binding to the groove in double-stranded DNA (blue). The six amino acids (residues) most commonly mutated in p53 are shown in yellow (from Cho et al., 1994).

So that was how P53 ‘talked’ to DNA to control the expression of specific genes. What could be better then, in a talk on how DNA damage can lead to cancer, than the story of a specific chemical doing nasty things to a gene that encodes perhaps the most revered of anti-cancer proteins?

The only thing baffling the students must have been the tobacco companies insisting, as they continued to do for years, that smoking was good for you.

And twenty-something years on …?

Well, it’s taken a couple of revolutions (scientific, of course!) but in that time we’ve advanced to being able to sequence genomes at a fantastic speed for next to nothing in terms of cost. In that period too more and more data have accumulated showing the pervasive influence of the weed. In particular that not only does it cause cancer in tissues directly exposed to cigarette smoke (lung, oesophagus, larynx, mouth and throat) but it also promotes cancers in places that never see inhaled smoke: kidney, bladder, liver, pancreas, stomach, cervix, colon, rectum and white blood cells (acute myeloid leukemia). However, up until now we’ve had very little idea of what, if anything, these effects have in common in terms of molecular damage.

Applying the power of modern sequencing, Ludmil Alexandrov of the Los Alamos National Lab, along with the Wellcome Trust Sanger Institute’s Michael Stratton and their colleagues have pieced together whole-genome sequences and exome sequences (those are just the DNA that encode proteins – about 1% of the total) of over 5,000 tumours. These covered 17 smoking-associated forms of cancer and permitted comparison of tobacco smokers with never-smokers.

Let’s hear it for consistent science!

The most obvious question then is do the latest results confirm the efforts of Denissenko & Co., now some 20 years old? The latest work found that smoking could increase the mutation load in the form of multiple, distinct ‘mutational signatures’, each contributing to different extents in different cancers. And indeed in lung and larynx tumours they found the guanine-to-thymine base-pair change that Denissenko et al had observed as the result of a specific chemical attaching to DNA.

For lung cancer they concluded that, all told, about 150 mutations accumulate in a given lung cell as a result of smoking a pack of cigarettes a day for a year.

Turning to tissues that are not directly exposed to smoke, things are a bit less clear. In liver and kidney cancers smokers have a bigger load of mutations than non-smokers (as in the lung). However, and somewhat surprisingly, in other smoking-associated cancer types there were no clear differences. And even odder, there was no difference in the methylation of DNA between smokers and non-smokers – that’s the chemical tags that can be added to DNA to tune the process of transforming the genetic code into proteins. Which was strange because we know that such ‘epigenetic’ changes can occur in response to external factors, e.g., diet.

What’s going on?

Not clear beyond the clear fact that tissues directly exposed to smoke accumulate cancer-driving mutations – and the longer the exposure the bigger the burden. For tissues that don’t see smoke its effect must be indirect. A possible way for this to happen would be for smoke to cause mild inflammation that in turn causes chemical signals to be released into the circulation that in turn affect how efficiently cells repair damage to their DNA.

raleighs_first_pipe_in_england-jpeg

Sir Walt showing off on his return                         to England

Whose fault it is anyway?

So tobacco-promoted cancers still retain some of their molecular mystery as well as presenting an appalling and globally growing problem. These days a popular pastime is to find someone else to blame for anything and everything – and in the case of smoking we all know who the front-runner is. But although Sir Walter Raleigh brought tobacco to Europe (in 1578), it had clearly been in use by American natives long before he turned up and, going in the opposite direction (à la Marco Polo), the Chinese had been at it since at least the early 1500s. To its credit, China had an anti-smoking movement by 1639, during the Ming Dynasty. One of their Emperors decreed that tobacco addicts be executed and the Qing Emperor Kangxi went a step further by beheading anyone who even possessed tobacco.

And paying the price

And paying the price

If you’re thinking maybe we should get a touch more Draconian in our anti-smoking measures, it’s worth pointing out that the Chinese model hasn’t worked out too well so far. China’s currently heading for three million cancer deaths annually. About 400,000 of these are from lung cancer and the smoking trends mean this figure will be 700,000 annual deaths by 2020. The global cancer map is a great way to keep up with the stats of both lung cancer and the rest – though it’s not for those of a nervous disposition!

References

Denissenko, M.F. et al. ( (1996). Preferential Formation of Benzo[a]pyrene Adducts at Lung Cancer Mutational Hotspots in P53.Science 274, 430–432.

Cho, Y. et al. (1994). Crystal Structure of a p53 Tumor Suppressor-DNA Complex: Understanding Tumorigenic Mutations. Science, 265, 346-355.

Alexandrov, L.D. et al. (2016). Mutational signatures associated with tobacco smoking in human cancer. Science 354, 618-622.

Seeing a New World

May I wish readers a Happy New Year – and indeed extend my felicitations to non-readers with the hope that they too will become followers! What a good idea! Not least because I suspect many are viewing the new year with a mixture of anxiety and despair. But I can promise there’s nothing like the sanity of science to restore you after a few minutes contemplating how we’re doing on the economic and political fronts.

Your starter for 2017

By happy chance a few weeks ago I tried to explain how it’s now possible to ‘re-write the manual of life’ – that is, to engineer our DNA, to fix broken genes if you like. This means that, in theory, it’s possible to correct errors in our genetic code that cause genetic diseases. As there are over 6,000 of these and they include Down syndrome, cystic fibrosis and Alzheimer’s disease, there’s no need to say it’s important. There are several ways of going about this but the one I described is called CRISPR and it’s had a lot of media coverage.

Right on cue

Well done then Keiichiro Suzuki, Juan Carlos Belmonte and friends from the Salk Institute in California together with colleagues from other centres in Spain, Saudi Arabia and China for their December paper describing a new CRISPR twist. They used a rat model of retinitis pigmentosa, a genetic disease that is a major cause of inherited blindness, afflicting about one and a half million people worldwide (one in 4,000 in the UK).

The CRISPR-Cas9 system is great but it works best in dividing cells (e.g., in skin and gut that are renewing all the time) and it’s particularly useful for knocking out genes rather than inserting new DNA. The latest modification allows a new gene to be inserted into a specific site in the DNA of cells that are not dividing (e.g., those of the eye or brain).

The bits of CRISPR-Cas9, which insert DNA at very precise locations within the genome, are delivered to target cells as part of an inert virus. However, the package also includes DNA that encourages the cells to use a repair process that can be turned on even in non-dividing cells. So CRISPR-Cas9 cuts the cell’s DNA at an exact sequence and the cell then repairs the double-strand breaks (by a process called non-homologous end joining (NHEJ) that glues the broken ends directly together). Give the cell a new bit of DNA (e.g., your favorite gene) and that will get patched in – bear in mind that the cell doesn’t ‘know’ what it’s doing: it just tries to fix damaged DNA with whatever’s at hand.

And the target?

Retinitis pigmentosa occurs when a chunk of a gene called Mertk is lost. After quite a lot of experiments to show that their method worked, Suzuki, Belmonte & Co made a viral carrier that included a normal Mertk gene and injected it under the retina of rats with the disease. After about 5 weeks the rats were making Mertk RNA as a result of the gene being correctly ‘knocked-in’ to eye cells. The light-detecting region of the eye, greatly reduced by the disease, was significantly restored, with associated appearance of MERTK protein.

      Diseased    Normal     Treated                         Diseased         Normal         Treated

pic

Left trio: Sections of the light-detecting layers of the eye in diseased (left), normal (centre) and diseased post-treatment rats (right). Right trio: corresponding fluorescence images showing MERTK expression (red: highlighted by white arrows); Cells labeled blue. (Suzuki et al. Nature 1–6 (2016) doi:10.1038/nature20565)

How did the rats see it?

Well, after treatment they were able to detect light and had significantly recovered their visual functions, albeit not to completely normal levels.

The usual caveats apply: the method isn’t hyper-efficient and a human treatment is still a long way off. Nevertheless, it’s a significant step.

The same group has also shown, using a way of re-programming the expression of just four genes, that it’s possible to arrest the signs of ageing. In other words, in mice this time, tinkering with these genes can increase lifespan – and yes, we have versions of these genes and in us they also control cell renewal.

So the New Year message is clear to see. If we can avoid turning the planet into a desert or blowing ourselves to smithereens the future is really rosy – and maybe even infinite!

References

Suzuki, K. et al. (2016). In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration. Nature 540, 144-149.

Ocampo, A. et al. (2016). In Vivo Amelioration of Age-Associated Hallmarks by Partial Reprogramming. Cell 167, 1719–1733.

Cockles and Mussels, Alive, Alive-O!

And so they are across the globe, not forgetting clams, a term that can cover all bivalve molluscs – a huge number of species (over 15,000), all having a two-part, hinged shell. The body inside doesn’t have a backbone, making it soft and edible on a scale of keeping-you-alive to orgasmic, depending on the consumer – oysters and scallops are part of the family.

Bivalves are particularly common on rocky and sandy coasts where they potter happily along, generally burrowing into sediment although some of them, scallops for instance, can swim. By and large their only problem is that humans like to eat them.

Clamming up

However, it gradually emerged in the 1970s that there was another cloud hovering over some of these gastronomic delights. Their commercial importance had drawn attention to the fact that soft-shell clams living along the east coast of North America, together with mussels on the west coast and cockles in Ireland, were dying in large numbers. The cause was an unusual type of cancer in which leukemia-like cells reproduce until they turn the blood milky and the animals die, in effect, from asphyxiation. In soft-shell clams, also known as sand gapers and steamers, the disease has spread over 1,500 km from Chesapeake Bay to Prince Edward Island.

A 2009 study had shown that as the disease progresses there is a rise in the number of blood cells that have abnormally high amounts of DNA (in clams typically four times the normal number of chromosomes – i.e. they’re tetraploid). In parallel with this change the cells make increasing amounts of an enzyme called reverse transcriptase (RT).

That was pretty surprising as RT does what its name suggests: reverses part of the central dogma of molecular biology (DNA makes RNA makes protein) by using RNA as a template to make DNA. RT is usually carried by viruses whose hereditary material is RNA (rather than DNA – so they’re called retroviruses). As part of their life cycle they turn their genomes into DNA that inserts into the host’s genome – which gets reproduced (as RNA) to make more viruses.

But how did RT get into clams? Enter Michael Metzger and Stephen Goff from Columbia University in New York, together with Carol Reinisch and James Sherry from Environment Canada, who began to unravel the mystery.

Jumping genes

Using high throughput sequencing they showed that clam genomes contain stretches of about 5,000 bases that came about when the RNA of a virus was copied into DNA by RT (reverse transcriptase) and then inserted into the host chromosome. Normal clams have from two to ten copies of this ‘repetitive element’ that Metzger & Co dubbed Steamer. That wasn’t too surprising as we have repetitive DNA too – it makes up about half the human genome. Many of these repeated sequences can move around within the genome – they’re often called ‘jumping genes’ – and it’s easy to see how this can happen when RT uses RNA to make DNA that can then pop into new sites in the genome. And you might guess that this process could damage the host DNA in ways that might lead to disease.

A long jump?

It turned out that the diseased clams had suffered massive amplification of Steamer to the extent that they carry 150 to 300 copies of the sequence. So that’s about 30 times as many Steamer DNAs being scattered across the clam genome – but how could that cause the same disease all the way from New York to Prince Edward Island? The answer came from peering into the DNA sequences of the tumour cells: they were virtually identical to each other – but they were different to those of their hosts! Meaning? The damage that led to leukemia, caused by shoe-horning 100s of extra copies of Steamer into clam genomes, only occurred once. And the staggering implication of that finding is that the cancer spread from a single ‘founder’ clam throughout these marine-dwelling molluscs. The resemblance to the way the cancer spreads in Tasmanian devils is striking.

Fishier and fishier

Fast forward to June 2016 and the latest contribution from the Metzger group reporting four more examples of transmissible cancer in bivalves – in mussels from British Columbia, in golden carpet shell clams from the Spanish coast and two forms in cockles.

Each appears to cause the same type of leukemia previously found in clams. The disease appears to be transmitted ‘horizontally’, i.e. by living cancer cells, descended from a single common ancestor, passing directly from one animal to another. Indeed, if you transplant blood cells from infected animals into normal clams they get leukemia.

 Species hopping

All that is quite amazing but the genetic analysis came up with an even more bizarre finding. In the golden carpet shell clams DNA from cancer cells showed no match with normal DNA from this species. It was clearly derived from a different species, which turned out to be the pullet shell clam – a species that, by and large doesn’t get cancer. So they have presumably come up with a way of resisting a cancer that arose in them, whilst at the same time being able to pass live tumour cells on to another species!!clam-transfer-pic

Cancer cell transmission between different species of shellfish. Cancer cells can arise in one species (pullet shell clams) that do not themselves develop leukemia but are able to pass live cells to another species (golden carpet shell clams) that do get leukemia (Metzger et al. 2016).

We have no idea how the cancer cells survive transfer. It seems most likely that they are taken up through the siphons that molluscs use for feeding, respiration, etc. and then somehow get across the walls of the respiratory/digestive systems. In the first step they would have to survive exposure to sea water which contains a lot more salt than cells are happy in. The ‘isotonic’ saline used in drips to infuse patients contains 0.9% salt whereas seawater, with 3.5%, is ‘hypertonic’ – cells put in a hypertonic solution will shrink as water is drawn out of the cell into the surrounding solution. Presumably the cells shrivel up a bit but some at least take this in their stride and recover to reproduce in their new host. Equally obscure is how a species can protect itself from a cancer that it can pass to another species.

These amazing findings throw a different light on the care-free underwater life depicted in Disney’s The Little Mermaid, in which the popular song ‘Under the Sea’ fails to mention floating cancer.

Can this happen to us?!!

Well, not as far as we know. But the fact that the known number of cancers that can be passed from one animal to another has now risen to nine does make you wonder. However, there’s no evidence that it happens in humans in anything like the normal course of events. There are examples of person-to-person transfer, notably during organ transplantation, and there is one recent case of cancerous cells from a tapeworm colonising a human host. But these are very rare, the latter occurring in a patient with a severely weakened immune system, and there is no example of spread beyond two people.

Phew! What a relief! So now we can concentrate on following developments both in Tasmania and beneath the waves in the hope that, not only can we go on satisfying our lust for clam bakes and chowders, but that these incredible creatures will reveal secrets that will benefit mankind.

References

AboElkhair, M. et al. (2009). Reverse transcriptase activity associated with haemic neoplasia in the soft-shell clam Mya arenaria. Diseases of Aquatic Organisms 84, 57-63.

Arriagada, G. et al. (2014). Activation of transcription and retrotransposition of a novel retroelement, Steamer, in neoplastic hemocytes of the mollusk Mya arenaria. PNAS 2014 111 (39) 14175-14180; published ahead of print September 8, 2014, doi:10.1073/pnas.1409945111.

Metzger, M.J. et al. (2015). Horizontal Transmission of Clonal Cancer Cells Causes Leukemia in Soft-Shell Clams. Cell 161, 255–263.

Metzger, M.J. et al. (2016). Widespread transmission of independent cancer lineages within multiple bivalve species. Nature 534, 705–709.

Muehlenbachs, A. et al. (2015). Malignant Transformation of Hymenolepis nana in a Human Host. N Engl J Med 2015; 373:1845-1852.

Another Peek At The Poor Little Devils

A couple of years ago (July 2014) I wrote a piece called Heir of the Dog that featured Tasmanian devils. The size of a small dog, these iconic little chaps are the largest meat-eating marsupials in the world. I’d run into them at The Lone Pine Koala Sanctuary in Brisbane where they’re keeping company with the dozy, furry tree-climbers as part of a programme to save them – the devils, that is – from extinction by cancer.

animal-fact-guide

A Tasmanian devil. Photo: Animal Fact Guide

1024px-sarcophilus_harrisii_taranna

 

 

 

 

 

 

 

imgresTheir problem comes from their inclination to bite one another, thereby directly passing on living cancer cells (causing devil facial tumour disease – DFTD). At that time the only other known example of transmissible cancer was a rare disease in dogs (canine transmissible venereal tumour – CTVT).

Genetic archaeology

DNA sequencing (i.e. whole genome analysis) had shown that the sexually transmitted dog disease probably arose thousands of years ago in a wolf or East Asian breed of dog and that the descendants of those cells are now present in infected dogs around the world.

The same approach applied to the Tasmanian devil showed that the cancer first arose in a female. Cells derived from that original tumour have subsequently spread through the Tasmanian population, the clone evolving (i.e. genetically diverging) over time. In contrast to the canine disease, DFTD is probably not more than 20 years old. Nevertheless, it spread through the wild population to the extent that the species was listed as endangered in 2008 by the International Union for Conservation of Nature.

Which is why a lot of effort is going into saving them, one approach being a number of breeding programmes in mainland Australia, with the aim of transferring uninfected animals to Tasmania.

One good turn …?

We’re all in favour of saving the little fellows, even if you probably wouldn’t want one as a pet. But, smelly and ferocious as he is, the Tasmanian devil is turning out to be remarkable in ways that suggest they might repay our efforts to keep them going. Things have moved apace down under with Greg Woods, Ruth Pye, Elizabeth Murchison, Andrew Storfer and colleagues from the Universities of Tasmania, Cambridge, Southampton and Washington State making some remarkable discoveries.

Infected animals do indeed develop the most unpleasant, large tumours that are virtually 100% fatal – to the extent that DFTD has wiped out 80% of Tasmanian devils in just 20 years. But some animals survive, even though models of the epidemiology say they shouldn’t. Andrew Storfer’s group asked how they pulled off this trick by looking for genetic changes in almost 300 devils. Quite amazingly, they found that even in a period as short as 20 years there were seven different genes that appeared to have changed (i.e. mutated) in response to selection imposed by the disease. Five of these genes encode proteins known to be associated with cancer risk or the immune system in other mammals, including humans. It seems that the mutations help their immune system to adapt so that it can recognize and destroy tumour cells.

In parallel with those studies, Greg Woods and his team now have a vaccine that looks promising early in trials – in other words a way of boosting natural immunity. We are only just beginning to find ways of giving the human immune system a helping hand – hence the burgeoning field of immunotherapy – so anything that works in another animal might give some useful pointers for us.

sick-tasTasmanian devil facial tumour disease.

This has killed 80% of the wild Australian animals in just a few decades.

Photograph: Menna Jones.

As if that wasn’t enough, a second strain of cancer has been found in a small group of male Tasmanian devils. It causes fatal facial tumours that look much the same as the first DFTD. However, it has a completely different genetic cause – so different in fact that it carries a Y chromosome, clear indication that the two forms of the disease arose by quite distinct mechanisms – which makes this marsupial the only species known to be affected by two types of transmissible of cancer.

Milk and human kindness

On top of all that some brave souls at Sydney University, Emma Peel and Menna Jones, decided in that way that scientists do, to collect some milk from the ferocious furries, just to see if it was interesting. Astonishingly the marsupial milk contained small proteins (peptides) that could kill a variety of bugs. They’re called cathelicidins and one of the things they can target is methicillin-resistant Staphylococcus aureus – MRSA – one of the dreaded ‘superbugs’ that are resistant to penicillin and other antibiotics. It’s not clear whether these peptides help to protect the devils from cancer but if that’s how turns out it might be incredibly important for us. As for their antibiotic potential, well, as it’s predicted that by 2050 superbugs will be killing one of us every three seconds you could say that opportunity beckons.

So that’s all incredibly exciting – and not just for the Tassie devils. ­ But another reason for returning to this story is that the devils have recently been joined by another example of extraordinary cancer transmission – and this one comes from the last place on the planet that you’d look for it ….

References

Murchison, E.P. et al. (2012). Genome Sequencing and Analysis of the Tasmanian Devil and Its Transmissible Cancer. Cell 148, 780–791.

Pye, R.J. et al. (2016). A second transmissible cancer in Tasmanian devils. Proceedings of the National Academy of Sciences USA, 113, 374–379.

Epstein, B. et al. (2016). Rapid evolutionary response to a transmissible cancer in Tasmanian devils. Nature Communications 7, Article number: 12684: doi:10.1038/ncomms12684.

Peel, E. et al. (2016). Cathelicidins in the Tasmanian devil (Sarcophilus harrisii). Scientific Reports 6, Article number: 35019. doi:10.1038/srep35019.

Long-live the Revolutions!!

There’s a general view that most folk don’t know much about science and, because almost day by day, science plays a more prominent role in our lives, that’s considered to be a Bad Thing. Us scientists are therefore always being told to get off our backsides and spread the word – and I try to do my bit in Betrayed by Nature, in Secret of Life (a new book shortly to be published) and in these follow-up blogs.

We may be making some progress – and, I have to admit, television has probably done more than me – though I am available (t.v. & movie head honchos please note). As one piece of evidence you could cite the way ‘DNA’ has become part of the universal lexicon, albeit often nonsensically. As evidence I call Sony Corp. Chief Executive Kazuo Hirai, as reported in The Wall Street Journal: “I’ve said this from day one. Some things at Sony are literally written into our DNA …”

Well, of course, that’s gibberish Kazuo old bean – but we know what you mean. Or do we? Most probably couldn’t tell you what the acronym stands for – but that doesn’t matter if they can explain that it’s the stuff (a ‘molecule’ would be better still!) that carries the information of inheritance and, as such, is responsible for all life. Go to the top of the class those who add that the code is in the form of chemicals called bases and there are just four of them (A, C, G & T). Something that simple doesn’t seem enough for all life but the secret is lies in the vast lengths of DNA involved. The human genome, for example, is made up of three billion letters.

A little bit of what is now history …

In the mid-1980s a number of scientists from around the world began to talk about the possibility of working out the sequence of letters that make up human DNA and thus identifying and mapping all the genes encoded by the human genome. From this emerged The Human Genome Project, a massive international collaboration, conceived in 1984 and completed in 2003. I quite often refer to this achievement as the ‘Greatest Revolution’ – meaning the biggest technical advance in the history of biology.

As that fantastic enterprise steadily advanced to its triumphant conclusion, it was accompanied by a series of mini-revolutions in technology that sky-rocketed the speed of sequencing and slashed the cost – the combined effect being an increase the efficiency of the whole process of more than 100 million-fold.

Brings us to the present …

These quite astonishing developments have continued since 2003 such that by 2009 it was possible to sequence 12 individuals in one study. By August 2016 groups from all over the world, coming together under the banner of The Exome Aggregation Consortium (ExAC), have raised the stakes 5,000-fold by sequencing no fewer than 60,706 individuals.

The name of the outfit tells you that there’s what you might think of as a very small swizz here: they didn’t sequence all the DNA, just the regions that code for proteins (exomes) – only about 1% of the three billion letters. But what highlights the power of current methods is not only the huge number of individuals sequenced but the depth of coverage – that is, the number of times each base (letter) in each individual exome was sequenced. In effect, it’s doing the same experiment so many times that errors are eliminated. Thus even genetic variants in just one person can be picked out.

seq-pic

Sequence variants between individuals. For most proteins the stretches of genomic DNA that encode their sequence  are split into regions called exons. All the expressed genes in a genome make up the exomeBy repeated sequencing The Exome Aggregation Consortium have shown that genetic variants in even one person can be reliably identified. Variants from the normal sequence found in four people are shown in red, bold letters.

It turns out that there are about 7.5 million variants and they pop up remarkably often – at one in every eight sites (bases). About half only occur once (which illustrates why DNA fingerprinting, aka DNA profiling, is so sensitive). As Jay Shendure put it, this gives us a “glimpse of the bottom of the well of genetic variation in humans.”

One of the major results of this study is that, by filtering out common variants from those associated with specific diseases, it will help to pin down the causes of Mendelian diseases (i.e. genetic disorders caused by change or alteration in a single gene, e.g., cystic fibrosis, haemophilia, sickle-cell anaemia, phenylketonuria). It’s clear that, over the next ten years, tens of millions of human genomes will be sequenced which will reveal the underlying causes of the thousands of genetic disorders.

The prize … and the puzzle

The technology is breathtaking, the amount of information being accumulated beyond comprehension. Needless to say, private enterprise has leapt on the bandwagon and you can now get your genome sequenced by, for example, 23andMe who offer “a personalised DNA service providing information and tools for individuals to learn about and explore their DNA. Find out if you are at risk for passing on an inherited condition, who you’re related to etc.” All for a mere $199!!

But you could say that the endpoint – the reason for grappling with DNA in the first place – is easy to see: eventually we will be able to define the molecular drivers of all genetic diseases and from that will follow ever improving methods of treatment and prevention.

Nevertheless, in that wonderful world I suspect we will still find ourselves brought up short by the underlying question: how one earth does DNA manage to carry the information necessary for all life?

For those who like to ponder such things, in the next piece we’ll try to help by looking at DNA from a different angle.

References

Ng, SB. et al. (2009). Nature 461, 272-276.

Lek, M. et al. (2016). Analysis of protein-coding genetic variation in 60,706 humans. Nature 536, 285–291.