Junk DNA: A Journey Through the Dark Matter of the Genome

5. Everything Shrinks When We Get Old

The movie Trading Places, starring Dan Aykroyd, Eddie Murphy and Jamie Lee Curtis, was a huge hit in 1983, grossing over $90 million at the US box office.^{44} It’s a convoluted comedy but the premise behind it is the exploration of genes versus environment. Is a successful man successful because of intrinsic merit or because of the environment in which he is placed? The movie comes out firmly on the side of the latter.

A similar phenomenon can happen in our genomes. An individual gene may perform a relatively innocuous role, helping a cell keep on keeping on, so to speak. The gene produces protein at just the right rate to do this job. A major factor in controlling the amount of protein that is produced is the position of the gene on the chromosome.

Now let’s imagine that the gene is transported to a new neighbourhood, like Dan Aykroyd’s character ending up in the slums or Eddie Murphy’s character finding himself transported to a mansion. In this neighbourhood, our transported gene is surrounded by new genomic information, which instructs it to make much higher amounts of protein. The high levels of the protein whip the cell forwards, pushing it to grow and divide much faster than usual. This can be one of the steps that leads to cancer. There’s nothing bad about the gene itself, it’s just in the wrong place at the wrong time.

This process is caused when two chromosomes break in a cell at the same time. When a chromosome breaks, a repair machinery immediately targets the break and joins the two bits up again. This is usually a pretty slick process. But if two (or more) chromosomes break at the same time, there can be problems. The ends of the chromosomes may become joined up incorrectly, as shown in Figure 5.1. This is how a ‘good’ gene may end up in a ‘bad’ neighbourhood, and begin causing problems. This is particularly an issue because the rearranged chromosomes will be passed on to all daughter cells every time cell division takes place. Probably the most famous example of this mechanism is in a human blood cancer called Burkitt’s lymphoma, where there is a rearrangement between chromosomes 8 and 14. This results in very strong over-expression of a gene[2] that encourages cells to proliferate aggressively.^{45}

Figure 5.1 In the upper panel a single chromosome breaks and is repaired by the cell. In the lower panel two chromosomes break simultaneously. The cell machinery may be unable to work out which break occurred on which chromosome. The chromosomes may be joined together inappropriately, creating hybrid structures.

Luckily, it’s probably quite rare that two chromosomes break at exactly the same time. More frequently there will be a time difference. So, the machinery that repairs DNA has evolved to act really quickly. After all, the faster it repairs a break, the lower the chance that there will be multiple breaks present at the same time in an individual cell. The DNA repair machinery starts to operate as soon as the cell detects that there is a broken piece of DNA. It does this by having mechanisms to detect the end of the break.

But this creates a whole new set of problems. Our cells contain 46 chromosomes, each of which is linear. In other words, our cells always have 92 chromosome ends, one at each end of a chromosome. The DNA damage machinery has to have a way of distinguishing the perfectly normal ends of chromosomes from the abnormal ends caused by breakages.

The way that cells have solved this is to have special structures on the normal ends of the chromosomes. Are you wearing shoes with laces? If so, have a quick look at those laces. At either end there is a little cap made from metal or plastic. This is called the aglet, and it stops the lace from unravelling and fraying. Our chromosomes have their own aglets, and these are extremely important for maintaining the integrity of our genome.

These chromosomal aglets are called telomeres and they are made from a form of junk DNA that we have known about for many years, plus complexes of various proteins. The telomeric DNA is formed from repeats of the same six base pairs, TTAGGG, repeated over and over again.^{46} These stretch for an average of about 10,000 base pairs in total on each end of every chromosome in the umbilical cord blood of a newborn human baby.^{47}

The telomeric DNA is bound by complexes of proteins that help to maintain the structural integrity.[3] The term telomere really refers to the combination of the junk DNA and its associated proteins. A graphic demonstration of the importance of these proteins was shown by some researchers working in mice in 2007. They knocked out expression of one of the proteins by completely inactivating its gene, and found that the resulting mice embryos died early in development.[4]

When the researchers examined the chromosomes in these genetically modified mice, they found that many of them had joined up. The ends had linked up with each other. This was because the DNA repair machinery no longer recognised the telomeres as telomeres. Instead, it reacted as if faced by a whole slew of broken chromosomes and did what it does best. It stuck them together. Unfortunately, by doing so, gene expression became completely disordered. Eventually the chromosomes and cells became so dysfunctional that they triggered a type of cellular suicide,[5] halting development completely.

There is also another feature of the telomeres that is of major interest in biology and human health. Back in the 1960s, researchers were studying how cells divide in the laboratory. They didn’t work with cancer cell lines, as these are derived from cells that have become immortal through abnormal changes. Instead, they studied a kind of cell known as a fibroblast. Fibroblasts are found in a wide range of human tissues. They secrete something called the extracellular matrix, a sort of thick wallpaper paste that holds the cells in position. It’s relatively easy to take a biopsy, for example from skin, and isolate the fibroblasts. These will grow and divide in culture. What the researchers discovered all those years ago was that the cells wouldn’t keep dividing forever. There came a point when they stopped dividing, even when supplied with all the nutrients and oxygen they needed. The cells didn’t die, they just stopped proliferating. This is known as senescence.^{48}

Scientists later realised that the telomeres in cells became shorter with each cell division. Every time one of the cells divided, all the DNA in that cell was copied. This ensured that both daughter cells inherited the same 46 chromosomes as the mother cell. But the system that copies the DNA in chromosomes can’t get right to the ends. So, over progressive cycles of cell division, the telomeres became shorter and shorter.^{49}

But this didn’t prove that the shortening of the telomeres actually caused cell senescence. It was perfectly possible that the effect on telomere length acted as a kind of marker for cell proliferation, but didn’t have any actual role to play in the changes in cell behaviour.

This is a really important concept in scientific enquiry. There are plenty of situations in which we can see a correlation between two things, but we shouldn’t from that automatically assume there is a causal relationship. Consider the following relationship. There is a strong relationship between developing lung cancer and sucking cough sweets. This doesn’t of course prove that sucking cough sweets gives you lung cancer. One of the first symptoms of lung cancer in many people is the development of a persistent cough, and someone with a cough is likely to try sucking hard sweets to decrease their discomfort.

The confirmation that telomere shortening did indeed lead to senescence came in the 1990s. Scientists demonstrated that if they increased the length of the telomeres in fibroblasts, the cells would bypass senescence and grow indefinitely.^{50}

It is now generally accepted that the telomeres act as a molecular clock, counting us down as we age. Not all the details have been established yet, because it’s a difficult area of biology to investigate, for a variety of reasons. One is that in any given cell, the 92 telomeric regions (one at each end of each chromosome) won’t be the same length. This makes it hard to come up with a meaningful measure of telomere length that is applicable throughout a cell, never mind an entire human being.^{51} It’s also very difficult for scientists to use their favourite model animal — the mouse — to investigate the relationships between telomere biology and ageing. This is because rodents have extremely long telomeres, much lengthier than in humans. Rodents, of course, are much shorter-lived than humans, suggesting that telomere length is not the only arbiter of ageing, but the accumulated evidence suggests that in humans they are of major importance.

What we do know is that our cells don’t succumb to the ageing process without a fight. They contain mechanisms to try to keep the telomeres long and intact as much as possible. This is achieved in our cells by something called telomerase activity. The telomerase system adds new TTAGGG motifs onto the ends of the chromosomes, basically restoring these important bits of junk DNA that are lost when the cells divide. Telomerase activity requires two components. One part is an enzyme, which adds the repeated sequences back on to the chromosome termini. The other is a piece of RNA, of a defined sequence, which acts as a template so that the enzyme adds the correct bases.

So the ends of our chromosomes rely heavily on junk DNA, genomic material that doesn’t code for proteins. The telomeres themselves are junk, and to maintain them the cell uses the output from a gene that produces RNA, but which is never used as a template for a protein. This RNA itself is a functional molecule, carrying out a vital role.[6]^,{52}

But if our cells contain a mechanism for maintaining telomere length, through the activity of the telomerase system, why do the telomeres get progressively shorter? What’s wrong with the system, why doesn’t it work properly?

The reason probably stems from the fact that there are few systems in biology that work well if allowed to run unchecked. And telomerase activity is held in very tight check indeed in our cells. The pathological exception to this is in cancer cells. Cancer cells frequently have adapted in such a way that they express high levels of telomerase activity and have elongated telomeres. This contributes to the aggressive growth and proliferation of many tumours. Our cellular systems have probably reached an evolutionary compromise. The telomeres are maintained at sufficient levels that we live long enough to reproduce (anything after that is irrelevant in evolutionary terms). But they aren’t so long that we succumb to cancer too early.

The basic telomere length in an individual is set fairly early in development, at a time when there is an uncharacteristic spike in the telomerase activity.^{53} Telomerase activity is also high in germ cells, the cells that give rise to eggs and sperm.^{54} This is to ensure that our offspring inherit telomeres of a good length.

Many human tissues contain cells known as stem cells. These are responsible for producing replacement cells when needed. When new cells are needed, a stem cell will copy its DNA and then split it between two daughter cells. Typically, one of these daughter cells will develop into a fully fledged replacement cell. The other will become a new stem cell, which can continue to create replacements in the same way.

One of the ‘busiest’ cell types in the human body is the type of stem cell that gives rise to all the blood cells,[7] including red blood cells and those that we rely on to fight infection. These stem cells proliferate at an incredible rate. This is because we constantly need to replenish the immune cells that fight off the foreign pathogens we encounter every day of our lives. We also need to replace red blood cells, because these only survive for about four months. Incredibly, the human body produces about 2 million red blood cells every second.^{55} That requires an awfully active stem cell population, in a pretty much constant state of cell division. These stem cells are enriched for telomerase activity, but eventually even they suffer from telomeres that are too short to do their job properly.^{56},{57} This is one reason why the elderly are at greater risk of infection than younger adults. They are essentially running out of immune cells. It’s also one of the reasons why cancer rates rise with age. Our immune system usually does a good job of destroying abnormal cells, but the effectiveness of this surveillance declines as stem cells die off.

Why is the length of our telomeres so important? It’s only junk DNA, so why should it matter if there are only several hundred copies of the non-coding TTAGGG, rather than a few thousand? Much of the problem seems to lie in the relationship between the DNA at the telomeres and the protein complexes that are deposited on this DNA. If the repetitive DNA shrinks below a critical level, the end of the chromosome can’t bind enough of the protective proteins. We’ve already seen one of the consequences of a lack of the relevant proteins in the mice that died before birth.

That was a very extreme example, but it’s undoubtedly the case that it’s vital that the telomeres are long enough to bind lots of the protective protein complexes. We know that this is true in humans as well as mice, because there are people who have inherited mutations in certain key components of the systems for maintaining the telomeres. The effects witnessed aren’t as dramatic as in the genetically modified mice, but that’s because such severely affected foetuses will tend to be lost during pregnancy. But the mutations we know about lead to conditions associated with certain disorders that are normally age-related.

The disorders are predominantly caused by mutations in the telomerase gene, or in the gene that codes for the RNA template, or in genes that encode proteins that protect the telomeres, or help the telomerase system to work effectively.[8]

Essentially, mutations in any of these genes can have similar effects. They basically make it harder for cells to maintain their telomeres. Consequently, the telomeres in patients with these mutations shorten more rapidly than in healthy individuals. This is why they develop symptoms that are suggestive of premature ageing. These disorders are known as human telomere syndromes.^{58}

Dyskeratosis congenita is a rare genetic condition, affecting about one in a million individuals. Patients suffer from a whole raft of problems. Their skin contains random dark patches. They develop white patches in their mouth, which can progress to oral cancer, and their fingernails and toenails are thin and weak. They suffer progressive and seemingly irreversible organ failure, triggered initially by bone marrow failure and lung problems. They are also at increased risk of cancer.

Scientists have realised that this condition can be caused by mutations in different genes in different affected families. At least eight mutated genes are known at the moment, and it’s quite possible that there are more.^{59} The feature that all the genes have in common is that they are involved in maintaining telomeres. This shows us that no matter how this region of junk DNA gets messed up, the final symptoms tend to be similar.

The lung problems are known as pulmonary fibrosis. Patients suffering from this condition have debilitating symptoms. They suffer shortness of breath and cough a lot, because they can’t move carbon dioxide out of their lungs efficiently or get oxygen into them easily. Looking at their lungs down a microscope, pathologists can see substantial regions where the normal tissue has been replaced by inflammation and fibrous tissue, rather like scar formation.^{60}

These clinical and pathological findings in the lungs are ones that are seen quite commonly in respiratory disease, and this prompted scientists to look at samples from patients with a condition known as idiopathic pulmonary fibrosis. Idiopathic just means that there is no obvious reason for the disease. Researchers tested these patients to see if any of them also had defects in the genes whose products protect the telomeres. In all, up to one in six people with a family history of this disease, but no previously identified mutations, were shown to have defects in the relevant genes.^{61},{62} Even in patients where there was no apparent family history of pulmonary fibrosis, mutations in telomere-relevant genes were found in between 1 and 3 per cent of cases.^{63},{64} There are about 100,000 patients with idiopathic pulmonary fibrosis in the United States, so at a conservative estimate 15,000 of them probably have developed the disease because they cannot maintain their telomeres properly.

Defects in the mechanisms that protect telomeres can also cause a different disease. There’s a condition called aplastic anaemia, in which the bone marrow fails to produce enough blood cells.^{65} It’s rare, affecting about one person in half a million. About one in twenty of the people with this condition have mutations in the telomerase enzyme or the accessory RNA template.

What may be happening in some of these patients is that they have both bone marrow defects and lung defects, but one problem becomes clinically apparent before the other. This can lead to unexpected consequences when medically treated. Bone marrow transplants are one of the treatments used for patients with aplastic anaemia. The patients are given drugs to prevent their immune system from rejecting the new bone marrow. Some of these drugs are known to have toxic effects in the lungs. For most patients with aplastic anaemia, this isn’t really a problem. But for those patients who have defects in their telomerase system, these drugs can trigger lung fibrosis that may actually be lethal.^{66} The cure becomes the cause of death.

There’s an odd genetic reason why clinicians may not realise that the symptoms they see in a patient are part of an inherited telomere problem. The telomerase complex is usually active in the germ cells, so that parents pass on long telomeres to their children. But in some of the families where there are mutations in the genes encoding the telomerase enzyme or the accessory RNA factor, this isn’t the case. As a consequence, each generation passes on shorter telomeres to its offspring. Because symptoms develop when the telomeres fall below a certain length, each successive generation is born rather nearer to the point where their telomere length falls over the cliff edge.^{67}

The effects of this are quite dramatic. A grandparent may have relatively long telomeres and develop pulmonary fibrosis in their 60s. Their child may have intermediate-length telomeres and develop lung symptoms in their 40s. But the third generation may inherit really short telomeres. They may develop aplastic anaemia in childhood.

Because the grandparental and parental generations’ conditions don’t develop until quite late in life, the grandchild may become sick before any of its elders have started displaying symptoms. This will make it difficult for a clinician to recognise that a genetic disease is present in the family, and this is compounded by the different symptoms found in the most severely and least severely affected individuals.

This strange pattern, where the oldest generation has different and milder symptoms that develop later in life than those found in the youngest generation, is rather similar to the inheritance pattern we saw in Chapter 1 for myotonic dystrophy. This is a very unusual genetic phenomenon and it is striking that in the two most clear-cut examples of this, the effect is ultimately caused by a change in length of a stretch of junk DNA.

One obvious question is why some tissues are more susceptible to short telomeres than others. This isn’t altogether clear, but some interesting models are emerging. It’s likely that tissues where there is a lot of proliferation will be susceptible to defects that lead to shorter telomeres. The classic example is the blood stem cell population, as described earlier in this chapter. If these cells have difficulties maintaining the length of their telomeres then eventually the stem cell population will run out.

That seems like a possible explanation for aplastic anaemia but it won’t work for pulmonary fibrosis. Lung tissue replicates quite slowly, yet pulmonary fibrosis is common in people with telomere defects. It’s possible that in lung cells the effects of shortened telomeres operate in tandem with other factors that affect the genome and cell function. These take time to develop, so lung symptoms typically develop later than ones that are caused by problems with the blood stem cells.

Our lungs are exposed to potentially damaging chemicals with every breath we take, so perhaps it’s not surprising that they struggle to tolerate the burden of defective telomeres. One of the most common sources of dangerous inhaled chemicals is tobacco. The global impact of smoking tobacco on human health is huge. The World Health Organization estimates that nearly 6 million people die every year as a consequence of smoking, over half a million of them from the effects of second-hand smoke.^{68}

Researchers examined the effects of cigarette smoke experimentally. They genetically manipulated mice so that some of them had short telomeres and then exposed various mice to cigarette smoke.^{69} The results are shown in Figure 5.2. Essentially, the only mice that developed pulmonary fibrosis were those that had short telomeres and were exposed to cigarette smoke.

Figure 5.2 A genetic defect and an environmental challenge are required to produce pulmonary fibrosis in mice. Mice with shortened telomeres don’t develop fibrosis, and nor do mice exposed to cigarette smoke. But mice with the double insult of shortened telomeres and exposure to cigarette smoke do develop the condition.

Cigarette smoking is not the only factor that affects human health, of course, although not smoking is probably the single smartest thing you can do for yourself. But the major factor that affects human health in wealthy countries is age itself. This wasn’t always the case. But it has been true since we made giant medical, pharmacological, social and technological progress in combating what used to kill us early: all those old-fashioned things like infectious diseases, early childhood mortality and malnutrition.

Getting old is now the major risk factor for development of chronic conditions. That’s a big problem when we realise that by 2025 there are likely to be over 1.2 billion people above the age of 60 worldwide.^{70} Cancer rates rise dramatically over the age of 40. If you live to 80, there’s an even chance you will develop some type of cancer. If you are over 65 and you’re an American, there’s about the same chance you will have cardiovascular disease.^{71} There’s plenty more statistics that paint a similarly bleak portrait, but why depress ourselves? Oh what the heck, one last one: the Royal College of Psychiatrists in the UK has stated that about 3 per cent of over-65s have clinical depression and one in six has symptoms of milder depression that are noticed by others.^{72}

Yet we all know that two individuals of the same chronological age may be very different in their health. Steve Jobs, the co-founder of Apple, died from cancer at the age of 56. Fauja Singh ran his first marathon at the age of 89, and his last at the age of 101 (no, it wasn’t the same one). There’s a lot we don’t know about what controls longevity — it is almost always a combination of genetics, environment and sheer luck. But what we do know is that simply counting how many years someone has been alive only gives you a very partial picture.

We are starting to realise that telomeres may be quite a sophisticated molecular clock. The rate of telomere shortening can be influenced by environmental factors. This means we may be able to use them as markers not of simple chronology, but of healthy years. The data are rather preliminary and not always consistent. This is partly because measuring telomeres in a consistent way is challenging, as described earlier, and we usually measure them in cells that we can access easily. These are typically the white blood cells, and they may not always be the most relevant cell type to examine. But despite these caveats, some intriguing data are emerging.

Let’s go back to our old enemy, tobacco. One study analysed the length of telomeres in the white blood cells of over 1,000 women. They found that the telomeres were shorter in those who smoked, with an increased rate of loss of about 18 per cent for every year of smoking. They calculated that smoking 20 cigarettes a day for 40 years was equivalent to losing almost seven and a half years of telomere life.^{73}

A 2003 study looking at mortality rates in the over-60s claimed that the people with the shortest telomeres had the highest mortality rates.^{74} This was mainly driven by cardiovascular mortality and the findings have been supported by a later, larger study in a different elderly population.^{75} A study in a group of centenarians from the Ashkenazi Jewish community found that longer telomeres were associated with fewer symptoms of the diseases of ageing, and with better cognitive function than that found in people of a similar advanced age but with shorter telomeres.^{76}

Sometimes we forget that it’s not just physical factors that affect health and longevity. Chronic psychological stress can be very harmful for an individual, with negative impacts on multiple systems including their cardiovascular health and their immune responses.^{77} Individuals who suffer chronic psychological stress tend to die younger than less stressed individuals. A study of women aged between 20 and 50 showed that those in the chronically stressed group had shorter telomeres than the unstressed women. This was calculated to equate to about ten years of life.^{78}

In the great pantheon of global human health problems that are eminently avoidable but having terrible impact, obesity seems to be on a mission to duke it out with smoking. Turning again to the World Health Organization we learn that nearly 3 million adults die each year because of being obese or overweight. Nearly a quarter of the burden of heart disease is attributable to people being overweight or obese. For type 2 diabetes, the contribution of obesity is even worse (almost half of all cases are caused by being overweight) and it’s also true for a significant proportion of cancers (between 7 and 41 per cent).^{79} The economic and social costs of this global epidemic are frightening.

Recent data have shown that there is significant interaction between the systems in our cells that try to regulate and respond to energy and metabolism fluctuations, and those that maintain genomic integrity, including telomere stability.^{80} It’s unsurprising, therefore, that scientists have analysed the lengths of telomeres in cells from obese individuals. The same paper that examined the effects of smoking on telomere length also looked at the effects of obesity. They found that the telomere shortening associated with obesity was even more pronounced than for smoking, equating to nearly nine years of life.^{81}

If all this inspires you to keep your weight under control, choose how you do this rather carefully. According to the United Nations, the country with the highest percentage of people who are aged 100 or over is Japan.^{82} The traditional Japanese diet almost certainly plays a role in this, because Japanese people who have changed to a Western diet develop Western chronic diseases. The traditional diet is based on low protein intake and relatively high carbohydrate levels. Studies in rats also showed that a low-protein diet early in life was associated with increased lifespan, which in turn was associated with long telomeres.^{83}

So if you’re thinking of adopting the high-protein and low-carb Atkins or Dukan diets, have a little word with your junk DNA first. I suspect your telomeres might say no.