Loss of Sirtuin Protein Seen to Cut Life Short

Evidence in Mice Connects Aging and DNA Repair

When the protein Sir2 was shown to extend life span in yeast, scientists were quick to hunt for genes in mammals that would do the same thing. But in mammals, the story is more complicated. Mammals have seven genes that are homologues of Sir2, called sirtuins. Only one of the proteins, SIRT1, has been studied in detail. Now a paper from the lab of Frederick Alt, a Howard Hughes investigator and the Charles A. Janeway professor of pediatrics at Children’s Hospital Boston, offers the first description of how a second member of the sirtuin family, SIRT6, functions in vivo. In the Jan. 27 Cell, Alt’s team describes how mice that lack SIRT6 display signs of premature aging and metabolic defects. The protein seems to play a role in stabilizing DNA, reminiscent of Sir2 in lower organisms.

Photo by Graham Ramsay

A second member of the sirtuin family, SIRT6, has some intriguing connections to aging, DNA repair, and metabolism that were uncovered by (left to right) Raul Mostoslavsky, Gustavo Mostoslavsky, Joyce Hsu, David Lombard, and Frederick Alt.

Researchers who study aging have been eager to see if the sirtuins hold the key to longevity in mammals. Alt, who is also the scientific director of the CBR Institute for Biomedical Research, did not set out to study aging; his lab was interested in sirtuins because they were thought to regulate DNA recombination. Because the lab already specialized in working with mice, the researchers became the first group to generate knockouts of all seven sirtuin genes.

Signs of Aging
After reporting results from their SIRT1 knockouts, Alt chose to focus on SIRT6, since mice lacking the gene showed the most obvious abnormalities after those lacking SIRT1. The team found that mice without SIRT6 appeared small at birth, but otherwise normal. After only 18 days, though, they rapidly degenerated, dying at four weeks. Their fat disappeared, their bones were weakened, their white blood cells diminished, their spleen and thymus vanished, and they suffered metabolic problems. “This doesn’t resemble a known disease,” said Raul Mostoslavsky, research fellow in genetics in Alt’s lab. Furthermore, each mouse enacted the same catastrophic sequence—there was very little individual variation in timing or symptoms, which usually occurs in illness.

Alt and Mostoslavsky are cautious about calling this phenomenon aging, but the symptoms matched many of the established signs of premature aging seen in other animal models.

Gene Guardian
Like Sir2, SIRT6 is found at the bundled chromatin inside a cell’s nucleus. Alt’s team discovered that the protein may work to keep DNA stable, but in a way different from Sir2’s. Cells lacking SIRT6 grew more slowly than control cells and were more sensitive to certain DNA-damaging agents. The chromosomes of these cells were laced with gaps and abnormalities.

Image courtesy of Raul Mostoslavsky Among their many signs of degeneration, SIRT6-deficient mice (right) experience bone loss and abnormal curvature of the spine after three weeks of age in comparison with normal mice, as shown in this X-ray.

Alt’s group, which included fellows Katrin Chua and David Lombard, collaborated with Bruce Demple, HSPH professor of toxicology and an expert in DNA repair. The team found that SIRT6-deficient cells were defective in a specific type of repair function, base excision repair, which is responsible for fixing spontaneous defects that crop up on a single strand of DNA. In this process, enzymes traveling along the DNA flip nucleotides out of place, scour them for defects, and substitute altered bases with sugar molecules; another set of enzymes then cuts these out and inserts new bases. This constant grooming of the genome is one of the cell’s primary defenses against oxidative DNA damage. Animals that lack key excision-repair proteins are not viable, but SIRT6 seems only to assist the process. The team found that adding a small active piece of polymerase beta, one of the key excision-repair components, could restore the cells’ ability to withstand damage.

For Demple, the finding is interesting because it provides “a possible connection between DNA repair and aging.” It has long been speculated that aging may result when the constant erosion of DNA by oxidative damage, a byproduct of normal metabolism, overcomes the cell’s ability to repair that damage.

Mostoslavsky said that because of its ability to stabilize DNA, SIRT6 may be the sirtuin that is closest functionally to Sir2 in yeast. SIRT1, the closest match genetically, removes acetyl groups from other proteins as Sir2 does. But instead of working on the level of DNA, where Sir2 helps to keep the genome stable, SIRT1’s primary function seems to lie elsewhere; it interacts with a host of other proteins, including the tumor suppressor p53 and insulin-signaling pathways. Several lines of research have shown that SIRT1 may mediate the extension of life span caused by calorie restriction.

Sirtuin Family History
The clinical state of the knockout mice suggests that SIRT6, like SIRT1, is also important in metabolism. The mice had severely low levels of the insulin-like growth factor and very low blood glucose, which the team speculates could have led to the demise of the animals’ immune systems. The team performed a bone marrow transplant from a SIRT6-deficient mouse to a normal mouse and found that the transplanted lymphocytes functioned normally. This suggests that the cells themselves are not lethally flawed; they “are responding to some kind of systemic defect,” Alt said.

David Sinclair, HMS associate professor of pathology, said that the study is an important step in filling in the sirtuin family picture. “This is really the first good evidence that the other members of the family are playing an important role in biology,” he said. “It’s not clear whether this is true premature aging, but it certainly does fit with the idea that sirtuins are longevity genes.”

So far, SIRT6 has yielded interesting links, but many unresolved questions. The findings dovetail with some of the most intriguing theories on the mechanisms of aging—metabolism and insulin signaling, DNA repair, and the accumulation of oxidative damage in cells. But like SIRT1, its function is undoubtedly more complex than its yeast counterpart. “We’re more complicated organisms; we don’t have one sirtuin, we have seven,” Alt said. “This family of molecules will be potentially very important,” but their importance may lie in unexpected areas, such as modifying metabolic disease. Alt’s lab is working to characterize the rest of the sirtuins to complete the family history.

—Courtney Humphries

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