Lifespan. Dr David A. Sinclair

lot of those things have to be put on hold.

      When sirtuins shift from their typical priorities to engage in DNA repair, their epigenetic function at home ends for a bit. Then, when the damage is fixed and they head back to home base, they get back to doing what they usually do: controlling genes and making sure the cell retains its identity and optimal function.

      But what happens when there’s one emergency after another to tend to? Hurricane after hurricane? Earthquake after earthquake? The repair crews are away from home a lot. The work they normally do piles up. The bills come due, then overdue, and then the folks from collections start calling. The grass grows too long, and soon the president of the neighborhood association is sending nastygrams. The baseball team goes coachless, and the team devolves into the Bad News Bears. And most of all, one of the most important things they do while at home—reproducing—doesn’t get done. This form of hormesis, the original survival circuit, works fine to keep organisms alive in the short term. But unlike longevity molecules that simply mimic hormesis by tweaking sirtuins, mTOR, or AMPK, sending out the troops on fake emergencies, these real emergencies create life-threatening damage.

      What could cause so many emergencies? DNA damage. And what causes that? Well, over time, life does. Malign chemicals. Radiation. Even normal DNA copying. These are the things that we’ve come to believe are the causes of aging, but there is a subtle but vital shift we have to make in that manner of thinking. It’s not so much that the sirtuins are overwhelmed, though they probably are when you are sunburned or get an X-ray; what’s happening every day is that the sirtuins and their coworkers that control the epigenome don’t always find their way back to their original gene stations after they are called away. It’s as if a few emergency workers who came to address the damage done in the Gulf Coast by Katrina had lost their home address. Then disaster strikes again and again, and they must redeploy.

      Wherever epigenetic factors leave the genome to address damage, genes that should be off, switch on and vice versa. Wherever they stop on the genome, they do the same, altering the epigenome in ways that were never intended when we were born.

      Cells lose their identity and malfunction. Chaos ensues. The chaos materializes as aging. This is the epigenetic noise that is at the heart of our unified theory.

      How does the SIR2 gene actually turn off genes? SIR2 codes for a specialized protein called a histone deacetylase, or HDAC, that enzymatically cleaves the acetyl chemical tags from histones, which, as you’ll recall, causes the DNA to bundle up, preventing it from being transcribed into RNA.

      When the Sir2 enzyme is sitting on the mating-type genes, they remain silent and the cell continues to mate and reproduce. But when a DNA break occurs, Sir2 is recruited to the break to remove the acetyl tags from the histones at the DNA break. This bundles up the histones to prevent the frayed DNA from being chewed back and to help recruit other repair proteins. Once the DNA repair is complete, most of the Sir2 protein goes back to the mating-type genes to silence them and restore fertility. That is, unless there is another emergency, such as the massive genome instability that occurs when ERCs accumulate in the nucleoli of old yeast cells.

      For the survival circuit to work and for it to cause aging, Sir2 and other epigenetic regulators must occur in “limiting amounts.” In other words, the cell doesn’t make enough Sir2 protein to simultaneously silence the mating-type genes and repair broken DNA; it has to shuttle Sir2 between the various places on an “as-needed” basis. This is why adding an extra copy of the SIR2 gene extends lifespan and delays infertility: cells have enough Sir2 to repair DNA breaks and enough Sir2 to silence the mating-type genes.³¹

      Over the past billion years, presumably millions of yeast cells have spontaneously mutated to make more Sir2, but they died out because they had no advantage over other yeast cells. Living for 28 divisions was no advantage over those that lived for 24 and, because Sir2 uses up energy, having more of the protein may have even been a disadvantage. In the lab, however, we don’t notice any disadvantage because the yeast are given more sugar than they could possibly ever eat. By adding 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