Lifespan

In yeast, the equivalent of the WRN gene is Slow Growth Suppressor 1, or SGS1. The gene was already suspected to code for a type of enzyme called a DNA helicase that untangles tangled strands of DNA before they break. Helicases are especially important in repetitive DNA sequences that are inherently prone to tangling and breaking. Functionality of proteins, such as the ones coded for by the Werner gene, is therefore vital, since more than half of our genome is, in fact, repetitive.

Through a gene-swapping process in which cells are tricked into picking up extra pieces of DNA, we swapped out the functional SGS1 gene with a mutant version. In effect, we were testing to see if it was possible to give the yeast Werner syndrome.

After the swap, the yeast cells’ lifespan was cut in half. Ordinarily, this would not have been news. Many events unrelated to aging—such as being eaten by a mite, drying out on a grape, or being placed in an oven—can and do shorten the lifespan of yeast cells. And here we’d messed with their DNA, which could have short-circuited the cells in a thousand different ways to cause early death.

But those cells weren’t just dying. They were dying after a precipitous decline in health and function. As the SGS1 mutants became older, they slowed down in their cell cycle. They grew larger. Both male and female “mating-type” genes (descendants of gene A) were switched on at the same time, so they were sterile and couldn’t mate. These were all known hallmarks of aging in yeast. And it was happening more quickly in the mutants we’d made. It certainly looked like a yeast version of Werner’s.

Using specialized stains, we colored the DNA blue and used red for the nucleolus, which sits inside the nucleus of all eukaryotic cells. That made it easier to see under the microscope what was happening at a cellular level.

And what was happening was fascinating.

The nucleolus is a part of the nucleus in which ribosomal DNA, or rDNA, resides. rDNA is copied into ribosomal RNA, which is used by ribosome enzymes to stitch amino acids together to make every new protein.

In the aged SGS1 cells, the nucleolus looked as if it had exploded. Instead of a single red crescent swimming in a blue ocean, the nucleolus was scattered into half a dozen small islands. It was tragic and beautiful. The picture, which would later appear in the August 1997 issue of the prestigious journal Science, still hangs in my office.

What happened next was both enchanting and illuminating. In response to the damage, like rats to the call of the Pied Piper, the protein called Sir2—the first known sirtuin, which is encoded by the gene SIR2[54 - SIR2 stands for “silent information regulator 2.” When SIR2 is written in capitals and italics, it refers to the gene; when it’s written Sir2, it refers to the protein.] and descended from gene B—had moved away from the mating genes that control fertility and into the nucleolus.

That was a beautiful sight to me, but it was a problem for the yeast. Sir2 has an important job: it is an epigenetic factor, an enzyme that sits on genes, bundles up the DNA, and keeps them silent. At the molecular level, Sir2 achieves this via its enzymatic activity, making sure that chemicals called acetyls don’t accumulate on the histones and loosen the DNA packaging.

When sirtuins left the mating genes—the ones descended from gene A that controlled fertility and reproduction—the mutant cells turned on both male and female genes, causing them to lose their sexual identity, just as in normal old cells, but much earlier.

I didn’t understand at first why the nucleolus was exploding, let alone why the sirtuins were moving toward it as the cells grew older. I agonized over the question for weeks.

And then one night, after a long day in the lab, I woke up with an idea.

It came in the space between sleep-deprived delirium and deep dreaming. The wisps of a concept. A few words jumbled together. A muddled picture of something. That was enough, though, to jolt me awake and pull me from my bed.

I grabbed my notebook and went to the kitchen. There, hunched over the table in the early morning hours of October 28, 1996, I began to write.

I wrote for about an hour, jotting down ideas, drawing pictures, sketching out graphs, formulating new equations.[55 - In a paper published in late 1997, I showed how ERCs—rDNA circles—cause aging and shorten the life of yeast cells. D. A. Sinclair and L. Guarente, “Extrachromosomal rDNA Circles—A Cause of Aging in Yeast,” Cell 91, no. 7 (December 26, 1997): 1033–42, https://www.ncbi.nlm.nih.gov/pubmed/9428525.] Scientific observations that had previously made no sense to me were falling perfectly into a larger picture. Broken DNA causes genome instability, I wrote, which distracts the Sir2 protein, which changes the epigenome, causing the cells to lose their identity and become sterile while they fixed the damage. Those were the analog scratches on the digital DVDs. Epigenetic changes cause aging.

There was, I imagined, a singular process that controlled them all. Not a countless number of separate cellular changes or diseases. Not even a set of hallmarks that could be addressed one at a time. There was something bigger—and more singular—than any of that.

This was the foundation for understanding the survival circuit and its role in aging.

The next day I showed Guarente my notes. I was excited; it felt like the biggest idea I’d ever had. But I was nervous, too; afraid he would find a hole in my logic and tear it apart. Instead, he looked over my notebook quietly, asked a few questions, and sent me on my way with six words.

“I like it,” he said. “Go prove it.”

THE RECITAL

To understand the Information Theory of Aging, we need to pay another visit to the epigenome, the part of the cell that the sirtuins help control.

Up close, the epigenome is more complex and wonderful than anything we humans have invented. It consists of strands of DNA wrapped around spooling proteins called histones, which are bound up into bigger loops called chromatin, which are bound up into even bigger loops called chromosomes.

Sirtuins instruct the histone spooling proteins to bind up DNA tightly, while they leave other regions to flail around. In this way, some genes stay silent, while others can be accessed by DNA-binding transcription factors that turn genes on.[56 - One way to think of the epigenome is as a cell’s software. In the same way digital files are stored in a phone’s memory and the software uses the ones and zeros to turn a phone into a clock, calendar, or music player, a cell’s information is stored as As, Ts, Gs, and Ts, and the epigenome uses those letters to direct a yeast cell to become male or a female and turn a mammalian cell into a nerve, a skin cell, or an egg.] Accessible genes are said to be in “euchromatin,” while silent genes are in “heterochromatin.” By removing chemical tags on histones, sirtuins help prevent transcription factors from binding to genes, converting euchromatin into heterochromatin.

Every one of our cells has the same DNA, of course, so what differentiates a nerve cell from a skin cell is the epigenome, the collective term for the control systems and cellular structures that tell the cell which genes should be turned on and which should remain off. And this, far more than our genes, is what actually controls much of our lives.

One of the best ways to visualize this is to think of our genome as a grand piano.[57 - I am not the first person to use this analogy. One of the earliest uses of the piano metaphor I can find came from a study guide intended to accompany a Nova Science NOW program on epigenetics in 2007. “Nova ScienceNOW: Epigenetics,” PBS, http://www.pbs.org/wgbh/nova/education/viewing/3411_02_nsn.html.] Each gene is a key. Each key produces a note. And from instrument to instrument, depending on the maker, the materials, and the circumstances of manufacturing, each will sound a bit different, even if played the exact same way. These are our genes. We have about 20,000 of them, give or take a few thousand.[58 - C. A. Makarewich and E. N. Olson, “Mining for Micropeptides,” Trends in Cell Biology 27, no. 9 (September 27, 2017): 685–96, https://www.ncbi.nlm.nih.gov/pubmed/28528987.]

Each key can also be played pianissimo (soft) or forte (with force). The notes can be tenuto (held) or allegretto (played quickly). For master pianists, there are hundreds of ways to play each individual key and endless ways to play the keys together, in chords and combinations that create music we know as jazz, ragtime, rock, reggae, waltzes, whatever.

The pianist that makes this happen is the epigenome. Through a process of revealing our DNA or bundling it up in tight protein packages, and by marking genes with chemical tags called methyls and acetyls composed of carbon, oxygen, and hydrogen, the epigenome uses our genome to make the music of our lives.

Yes, sometimes the size, shape, and condition of a piano dictate what a pianist can do with it. It’s tough to play a concerto on an eighteen-key toy piano, and it’s mighty hard to make beautiful music on an instrument that hasn’t been tuned in fifty years. Likewise, the genome certainly dictates what the epigenome can do. A caterpillar can’t become a human being, but it can become a butterfly by virtue of changes in epigenetic expression that occur during metamorphosis, even though its genome never changes. Similarly, the child of two parents from a long line of people with black hair and brown eyes isn’t likely to develop blond hair and blue eyes, but twin agouti mice in the lab can turn out brown or golden, depending on how much the Agouti gene is turned on during gestation by environmental influences on the epigenome, such as folic acid, vitamin B

, genistein from soy, or the toxin bisphenol A.[59 - D. C. Dolinoy, “The Agouti Mouse Model: An Epigenetic Biosensor for Nutritional and Environmental Alterations on the Fetal Epigenome,” Nutrition Reviews 66, suppl. 1 (August 2008): S7–11, https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2822875/.]

Similarly, among monozygotic human twins, epigenetic forces can drive two people with the same genome in vastly different directions. It can even cause them to age differently. You can see this clearly in side-by-side photographs of the faces of smoking and nonsmoking twins; their DNA is still largely the same, but the smokers have bigger bags under their eyes, deeper jowls below their chins, and more wrinkles around their eyes and mouths. They are not older, but they’ve clearly aged faster. Studies of identical twins place the genetic influences on longevity at between 10 and 25 percent which, by any estimation, is surprisingly low.[60 - The more extroverted you are, the longer your lifespan, while, perhaps unsurprisingly, pessimists and psychotics see significant increases in the risk of death at an earlier age. That’s according to a study of 3,752 twins 50 years or older that looked at the relationship between personality and lifespan through the prism of genetic influences. M. A. Mosing, S. E. Medland, A. McRae, et al., “Genetic Influences on Life Span and Its Relationship to Personality: A 16-Year Follow-up Study of a Sample of Aging Twins,” Psychosomatic Medicine 74, no. 1 (January 2012): 16–22, https://www.ncbi.nlm.nih.gov/pubmed/22155943. The authors considered definitions of extreme longevity, using multiple European twin registries. A. Skytthe, N. L. Pedersen, J. Kaprio, et al., “Longevity Studies in GenomEUtwin,” Twin Research 6, no. 5 (October 2003): 448–54, https://www.ncbi.nlm.nih.gov/pubmed/14624729.]

Our DNA is not our destiny.

Now imagine you’re in a concert hall. A virtuoso pianist is seated at a gorgeously polished Steinway grand. The concerto begins. The music is beautiful, breathtaking. Everything is perfect.

But then, a few minutes into the piece, the pianist misses a key. The first time it happens, it’s almost unnoticeable—an extra D, perhaps, in a chord that doesn’t need that note. Embedded in so many perfectly played notes, hidden among an otherwise flawless chord in an otherwise perfect melody, it’s nothing to worry about. But then, a few minutes later, it happens again. And then, with increasing frequency, again and again and again.

It’s important to remember that there is nothing wrong with the piano. And the pianist is playing most of the notes prescribed by the composer. She’s just also playing some extra notes. Initially, this is just annoying. Over time it becomes unsettling. Eventually it ruins the concerto. Indeed, we’d assume that there was something wrong with the pianist. Someone might even rush onto the stage to make sure she is all right.

Epigenetic noise causes the same kind of chaos. It is driven in large part by highly disruptive insults to the cell, such as broken DNA, as it was in the original survival circuit of M. superstes and in the old yeast cells that lost their fertility. And this, according to the Information Theory of Aging, is why we age. It’s why our hair grays. It’s why our skin wrinkles. It’s why our joints begin to ache. Moreover, it’s why each one of the hallmarks of aging occurs, from stem cell exhaustion and cellular senescence to mitochondrial dysfunction and rapid telomere shortening.

This is, I acknowledge, a bold theory. And the strength of a theory is based on how well it predicts the results of rigorous experiments, often millions of them, the number of phenomena it can explain, and its simplicity. The theory was simple, and it explained a lot. As good scientists, what we had left to do was to try our best to disprove it and see how long it survived.

To get started, Guarente and I had to get our eyes on some yeast DNA.

We used a technique called a Southern blot, a method of separating DNA based on its size and conformation and lighting it up with a radioactive DNA probe. In the first experiment, we noticed something spectacular. Normally, the rDNA of a yeast cell that is made visible by a Southern blot is tightly packed, like a new spool of rope, with a few barely visible wispy loops of supercoiled DNA. But the rDNA of the yeast cells we’d created in our lab—the Werner mutants that seemed to be aging rapidly—were madly unpacking, like a vacuum-sealed bag of yarn that had been ripped open.

LESSONS FROM YEAST CELLS ABOUT WHY WE AGE. In young yeast cells, male and female “mating-type information” (gene A) is kept in the “off” position by the Sir2 enzyme, the first sirtuin (encoded by a descendant of gene B). The highly repetitive ribosomal DNA (rDNA) is unstable, and toxic DNA circles form; these recombine and eventually accumulate to toxic levels in old cells, killing them. In response to DNA circles and the perceived genome instability, Sir2 moves away from silent mating-type genes to help stabilize the genome. Both male and female genes turn on, causing infertility, the main hallmark of yeast aging.

The rDNA was in a state of chaos. The genome, it seemed, was fragmenting. DNA was recombining and amplifying, showing up on the Southern blot as dark spots and wispy circles, depending on how coiled up and twisted they were. We called those loops extrachromosomal ribosomal DNA circles, or ERCs, and they were accumulating as the mutants aged.

If we had indeed induced aging, then we would see this same pattern emerge in yeast cells that had aged normally.

We don’t count the age of a single yeast cell with birthday candles. They simply don’t last that long. Instead, aging in yeast is measured by the number of times a mother cell divides to produce daughter cells. In most cases, a yeast cell gets to about 25 divisions before it dies. That, however, makes obtaining old yeast cells an exceptionally challenging task. Because by the time an average yeast cell expires, it is surrounded by 2

, or 33 million, of its descendants.

It took a week of work, a lot of sleepless nights, and a whole lot of caffeinated beverages to collect enough regular old cells. The next day, when I developed the film to visualize the rDNA, what I saw astounded me.[61 - It was a eureka moment—discovering why yeast cells age. Supercoiled circles of ribosomal DNA pinch off the yeast chromosome and accumulate as the yeast divide, distracting the Sir2 enzyme from its main role of controlling genes for sex and reproduction. David A. Sinclair and Leonard Guarente, “Extrachromosomal rDNA Circles—A Cause of Aging in Yeast,” Cell 91 (December 26, 1997): 1033–42.]

Just like the mutants, the normal old yeast cells were packed with ERCs.

That was a “Eureka!” moment. Not proof—a good scientist never has proof of anything—but the first substantial confirmation of a theory, the foundation upon which I and others would build more discoveries in the years to come.

The first testable prediction was if we put an ERC into very young yeast cells—and we devised a genetic trick to do that—the ERCs would multiply and distract the sirtuins, and the yeast cells would age prematurely, go sterile, and die young—and they did. We published that work in December 1997 in the scientific journal Cell, and the news broke around the world: “Scientists figured out a cause of aging.”

It was there and then that Matt Kaeberlein, a PhD student at the time, arrived at the lab. His first experiment was to insert an extra copy of SIR2 into the genome of yeast cells to see if it could stabilize the yeast genome and delay aging. When the extra SIR2 was added, ERCs were prevented, and he saw a 30 percent increase in the yeast cells’ lifespan, as we’d been hoping. Our hypothesis seemed to be standing up to scrutiny: the fundamental, upstream cause of sterility and aging in yeast was the inherent instability of the genome.

What emerged from those initial results in yeast, and another decade of pondering and probing mammalian cells, was a completely new way to understand aging, an information theory that would reconcile seemingly disparate factors of aging into one universal model of life and death. It looked like this: