Lifespan. David Sinclair
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extra copies of the SIR2 gene, we gave the yeast cells what evolution failed to provide.
If the information theory is correct—that aging is caused by overworked epigenetic signalers responding to cellular insult and damage—it doesn’t so much matter where the damage occurs. What matters is that it is being damaged and that sirtuins are rushing all over the place to address that damage, leaving their typical responsibilities and sometimes returning to other places along the genome where they are silencing genes that aren’t supposed to be silenced. This is the cellular equivalent of distracting the cellular pianist.
To prove that, we needed to break some mouse DNA.
It’s not hard to intentionally break DNA. You can do it with mechanical shearing. You can do it with chemotherapy. You can do it with X-rays.
But we needed to do it with precision—in a way that wouldn’t create mutations or impact regions that affect any cellular function. In essence, we needed to attack the wastelands of the genome. To do that, we got our hands on a gene similar to Cas9, the CRISPR gene-editing tool from bacteria that cuts DNA at precise places.
The enzyme we chose for our experiments comes from a goopy yellow slime mold called Physarum polycephalum, which literally means “many-headed slime.” Most scientists believe that this gene, called I-PpoI, is a parasite that serves only to copy itself. When it cuts the slime mold genome, another copy of I-PpoI is inserted. It is the epitome of a selfish gene.
That’s in a slime mold, its native habitat. But when I-PpoI finds itself in a mouse cell, it doesn’t have all the slime mold machinery to copy itself. So it floats around and cuts DNA at just a few places in the mouse genome, and there is no copying process. Instead, the cell has no problem pasting the DNA strands back together, leaving no mutations, which is exactly what we were looking for to engage the survival circuit and distract the sirtuins. DNA-editing genes such as Cas9 and I-PpoI are nature’s gifts to science.
To create a mouse to test the information theory, we inserted I-PpoI into a circular DNA molecule called a plasmid, along with all the regulatory DNA elements needed to control the gene, and then inserted that DNA into the genome of a mouse embryonic stem cell line we were culturing in plastic dishes in the lab. We then injected the genetically modified stem cells into a 90-cell mouse embryo called a blastocyst, implanted it into a female mouse’s uterus, and waited about twenty days for a baby mouse to show up.
This all sounds complicated, but it’s not. After some training, a college student can do it. It’s such a commodity these days, you can even order a mouse out of a catalog or pay a company to make you one to your specifications.
The baby mice were born perfectly normal, as expected, since the cutting enzyme was switched off at that stage. We called them affectionately “ICE mice,” ICE standing for “Inducible Changes to the Epigenome.” The “inducible” part of the acronym is vital—because there’s nothing different about these mice until we feed them a low dose of tamoxifen. This is an estrogen blocker that is normally used to treat human cancers, but in this case, we’d engineered the mouse so that tamoxifen would turn on the I-PpoI gene. The enzyme would go to work, cutting the genome and slightly overwhelming the survival circuit, without killing any cells. And since tamoxifen has a half-life of only a couple of days, removing it from the mice’s food would turn off the cutting.
The mice might have died. They might have grown tumors. Or they might have been perfectly fine, no worse off than if they’d received a dental X-ray. Nobody had ever done this before in a mouse, so we didn’t know. But if our hypothesis about epigenetic instability and aging was correct, the tamoxifen would work like the potion that Fred and George Weasley used to age themselves in Harry Potter and the Goblet of Fire.
And it worked. Like wizardry, it did.
During the treatment, the mice were fine, oblivious to the DNA cutting and sirtuin distraction. But a few months later, I got a call from a postdoc who was taking care of our lab’s animals while I was on a trip to my lab in Australia.
“One of the mice is really sick,” she said. “I think we need to put it down.”
I asked her to text me a photo of the mouse she was talking about. When the photo came over my phone, I couldn’t help but laugh.
“That’s not a sick mouse,” I replied. “That’s an old mouse.”
“David,” she said, “I think you’re mistaken. It says here that it’s the sister of these other mice in the cage, and they’re perfectly normal.”
Her confusion was understandable. At 16 months old, a regular lab mouse still has a thick coat of fur, a sturdy tail, a muscular figure, perky ears, and clear eyes. A tamoxifen-triggered ICE mouse at the same age has thinning, graying hair, a bent spine, paper-thin ears, and cloudy eyes.
Remember, we’d done nothing to change the genome. We’d simply broken the mice’s DNA in places where there aren’t any genes and forced the cell to paste, or “ligate,” them back together. Just to make sure, later we broke the DNA in other places, too, with the same results. Those breaks had induced a sirtuin response. When those fixers went to work, their absence from their normal duties and presence on other parts of the genome altered the ways in which lots of genes were being expressed at the wrong time.
THE MAKING OF THE ICE MOUSE TO TEST IF THE CAUSE OF AGING MIGHT BE INFORMATION LOSS. A gene from a slime mold that encodes an enzyme that cuts DNA at a specific place was inserted into a stem cell and injected into an embryo to generate the ICE mouse. Turning on the slime mold gene cut the DNA and distracted the sirtuins, causing the mouse to undergo aging.
Those findings were aligned to discoveries being made by Trey Ideker and Kang Zhang, at UC San Diego, and Steve Horvath, at UCLA. Steve’s name stuck, and today he’s the namesake of the Horvath Clock—an accurate way of estimating someone’s biological age by measuring thousands of epigenetic marks on the DNA, called methylation. We tend to think of aging as something that begins happening to us at midlife, because that’s when we start to see significant changes to our bodies. But Horvath’s clock begins ticking the moment we are born. Mice have an epigenetic clock, too. Were the ICE mice older than their siblings? Yes, they were—about 50 percent older.
We’d found life’s master clock winder.
In another manner of thinking, we’d scratched up the DVD of life about 50 percent faster than it normally gets scratched. The digital code that is, and was, the basic blueprint for our mice was the same as it had always been. But the analog machine built to read that code was able to pick up only bits and pieces of the data.
Here’s the vital takeaway: we could age mice without affecting any of the most commonly assumed causes of aging. We hadn’t made their cells mutate. We hadn’t touched their telomeres. We hadn’t messed with their mitochondria. We hadn’t directly exhausted their stem cells. Yet the ICE mice were suffering from a loss of body mass, mitochondria, and muscle strength and an increase in cataracts, arthritis, dementia, bone loss, and frailty.
All of the symptoms of aging—the conditions that push mice, like humans, farther toward the precipice of death—were being caused not by mutation but by the epigenetic changes that come as a result of DNA damage signals.
We hadn’t given the mice all of those ailments. We had given them aging.
And if you can give something, you can take it away.
FRUIT OF THE SAME TREE
Like the gnarled hands of giant zombies breaking free of the rocky soil, the ancient bristlecone pine trees of California’s White Mountains strike haunting silhouettes against the dewy morning sun.
The oldest of these trees have been here since before the pyramids of Egypt were built, before the construction of Stonehenge, and before the last of the woolly mammoths left our world. They have shared this planet with Moses, Jesus, Muhammad, and the first Buddha. Standing some two miles above sea level, adding fractions