Yeast have a specific feature that has made them one of the favourite model systems of epigeneticists. Yeast never methylate their DNA, so all epigenetic effects must be caused by histone modifications. There’s also another helpful feature of yeast. Each time a yeast mother cell gives rise to a daughter cell, the bud leaves a scar on the mother. This makes it really easy to work out how many times a cell has divided. There are two types of ageing in yeast and these each have parallels to human ageing, as shown in Figure 13.2.
Figure 13.2 The two models of ageing in yeast, relevant for dividing and non-dividing cells.
Most of the emphasis in ageing research has been on replicative ageing, and trying to understand why cells lose their ability to divide. Replicative ageing in mammals is clearly related to some obvious symptoms of getting older. For example, skeletal muscle contains specialised stem cells called satellite cells. These can only divide a certain number of times. Once they are exhausted, you can’t create new muscle fibres.
Substantial progress has been made in understanding replicative ageing in yeast. One of the key enzymes in controlling this process is called Sir2 and it’s an epigenetic protein. It affects replicative ageing in yeast through two pathways. One seems to be specific to yeast, but the other is found in numerous species right through the evolutionary tree, all the way up to humans.
Sir2 is a histone deacetylase. Mutant yeast that over-express Sir2 have a replicative lifespan that is at least 30 per cent longer than normal[241]. Conversely, yeast that don’t express Sir2 have a reduced lifespan[242], about 50 per cent shorter than usual. In 2009, Professor Shelley Berger, an incredibly dynamic scientist at the University of Pennsylvania whose group has been very influential in molecular epigenetics, published the results of a really elegant set of genetic and molecular experiments in yeast.
Her research showed that the Sir2 protein influences ageing by taking acetyl groups off histone proteins, and not through any other roles this enzyme might carry out[243]. This was a key experiment, because Sir2, like many histone deacetylases, has rather loose molecular morals. It doesn’t just remove acetyl groups from histone proteins. Sir2 will take acetyl groups away from at least 60 other proteins in the cell. Many of these proteins have nothing to do with chromatin or with gene expression. Shelley Berger’s work was crucial for demonstrating that Sir2 influences ageing precisely because of its effects on histone proteins. The altered epigenetic pattern on the histones in turn influenced gene expression.
These data, showing that epigenetic modifications of histones really do have a major influence on ageing, gave scientists in this field a big confidence boost that they were on the right track. The importance of Sir2 doesn’t seem to be restricted to yeast. If we over-express Sir2 in our favourite worm, C. elegans[244], the worm lives longer. Fruit flies that over-expressed Sir2 had up to a 57 per cent increase in lifespan[245]. So, could this gene also be important in human ageing?
There are seven versions of the Sir2 gene in mammals, called SIRT1 through to SIRT7. Much of the attention in the human field has focused on SIRT6, an unusual histone deacetylase. The breakthroughs in this field have come from the laboratory of Katrin Chua, a young Assistant Professor at the Stanford Center on Longevity (and also the sister of Amy Chua who wrote the highly controversial mothering memoir Battle Hymn of the Tiger Mother).
Katrin Chua created mice which never expressed any Sirt6 protein, even during their development (they are known as Sirt6 knockout mice). These animals seemed normal at birth, although they were rather small. But from two weeks of age onwards they developed a whole range of conditions that mimicked the ageing process. These included loss of fat under the skin, spinal curvature, and metabolism deficits. The mice died by one month of age, whereas a normal mouse can live for up to two years under laboratory conditions.
Most histone deacetylases are very promiscuous. By this we mean they will deacetylate any acetylated histone they can find. Indeed, as mentioned above, many don’t even restrict themselves to histones, and will take acetyl groups off all sorts of proteins. However, SIRT6 isn’t like this. It only takes the acetyl groups off two specific amino acids – lysine 9 and lysine 56, both on histone H3. The enzyme also seems to have a preference for histones that are positioned at telomeres. When Katrin Chua knocked out the SIRT6 gene in human cells, she found that the telomeres of these cells got damaged, and the chromosomes began to join up. The cells lost the ability to divide any further and pretty much shut down most of their activities[246].
This suggested that human cells need SIRT6 so that they can maintain the healthy structures of telomeres. But this wasn’t the only role of the SIRT6 protein. Acetylation of histone 3 at amino acid 9 is associated with gene expression. When SIRT6 removes this modification, this amino acid can be methylated by other enzymes present in the cell. Methylation at this position on the histone is associated with gene repression. Katrin Chua performed further experiments which confirmed that changing the expression levels of SIRT6 changed the expression of specific genes.
SIRT6 is targeted to specific genes by forming a complex with a particular protein. Once it’s present at those genes, SIRT6 takes part in a feedback loop that keeps driving down expression of the gene, in a classic vicious cycle. When the SIRT6 gene is knocked out, the levels of histone acetylation at these genes stays high because the feedback loop can’t be switched on. This drives up expression of these target genes in the SIRT6 knockout mice. The target genes are ones which promote auto-destruction, or the cell’s entry into a state of permanent stasis known as senescence. This effect explains why SIRT6 knockdown is associated with premature ageing[247]. It’s because genes that accelerate processes associated with ageing are switched on too soon, or too vigorously, at a young age.
It’s a little like a crafty manufacturer installing an inbuilt obsolescence mechanism into a product. Normally, the mechanism doesn’t kick in for a certain number of years, because if the obsolescence activates too early, the manufacturer will get a reputation for prematurely shoddy goods and nobody will buy them at all. Knocking out SIRT6 in cells is a little like a software glitch that activates the inbuilt obsolescence pathway after, say, one month instead of two years.
Other SIRT6 target genes are associated with provoking inflammatory and immune responses. This is also relevant to ageing, because some conditions that become much more common as we age are a result of increased activation of these pathways. These include certain aspects of cardiovascular disease and chronic conditions such as rheumatoid arthritis.
There is a rare genetic disease called Werner’s syndrome. Patients with this disorder age faster and at an earlier age than healthy individuals. The condition is caused by mutations in a gene that is involved in the three-dimensional structure of DNA, keeping it in the correct conformation and wound up to the right degree of tightness for a specific cell type[248]. The normal protein binds to telomeres. It binds most effectively when the histones at the telomeres have lost the acetyl group at amino acid 9 on histone H3. This is the precise modification removed by the SIRT6 enzyme. This further strengthens the case for a role of SIRT6 in control of ageing[249].
245
Rogina and Helfand (2004),