New & Noteworthy

Link Between Aging and Iron

August 25, 2022

Perturbations in iron homeostasis affect aging, but how this happens has remained a bit of a black box. A new study by Patnaik et al. in Cell Reports illuminates this box by looking more closely at the transcription factors that are first to respond when iron becomes limiting.

Key among these are Atf1p and Atf2p, which activate the full suite of iron-mobilization genes, among which is TIS11/CTH2, which encodes an RNA-binding protein that targets specific messages for decay.

The targeted messages flagged for decay encode mitochondrial proteins, as these use iron but are not the most essential in the set. The most essential Fe-requiring enzymes are those involved in DNA synthesis and repair, such that slowing/shutting mitochondrial function is a response to iron deficiency. Intriguingly, mitochondrial function also happens to decline with age.

To find the specific mechanisms linking iron with aging, the authors used an unbiased analysis of genes involved in iron homeostasis to see which showed connection with aging. The strain with a tis11Δ mutation lived longer than any others, with a lifespan extended by 51.1%. In a broader sense, they found that genes involved in different aspects of response to iron deficiency also had different effects on fitness and aging.

Delving more deeply into the role of Tis11p/Cth2p in aging, the authors used RNA-seq and Ribo-seq to look at temporal changes in transcription versus translation in aging cells. They showed how, overall, aging leads to inhibition of translation—except for certain genes which are upregulated instead. Interestingly, most of the upregulated genes are in the Fe regulon that gets activated by the first responder Atf1p.

While the expression of TIS11/CTH2 increases both with aging and with iron deficiency, the deletion of the gene extends lifespan. Thus, multiple lines of evidence suggest Tis11p/Cth2p is a negative regulator of longevity. The key connection appears to be mitochondrial translation, where the function of Tis11p/Cth2p to inhibit translation of mitochondrial transcripts for repressing non-essential Fe-requiring enzymes serves to simultaneously repress overall mitochondrial respiration, which speeds aging.

As not all genes translationally upregulated in the tis11Δ mutant contained appropriate binding sites in the 3’ UTR, the authors looked further and found binding sequences for Puf3p, a protein known to bind and inhibit translation of mRNAs coding for mitochondrial ribosome proteins. Thus, Puf3p appears to be a critical partner for Tis11p/Cth2p in mediating downregulation of mitochondrial function. Further, they questioned the relationship with the Hap4p transcription factor, which regulates numerous components of the electron transport chain and whose overexpression extends lifespan. As the combination of a tis11Δ deletion with HAP4 overexpression had no additive effect in an epistasis experiment, they concluded that Tis11p-dependent repression acts through Hap4p.

The role of phosphorylation of Tis11p/Cth2p was examined by mutating N-terminal serine residues, which impairs degradation of the protein. Consistent with the converse result of extended lifespan in null mutants, the nondegradable version of the protein shortens lifespan.

Thus, the ease of the yeast model once more illuminates intricate connections between critical proteins, facilitating potential drug discovery around several new aging factors.

Categories: Research Spotlight

Tags: aging, cell aging, iron homeostasis, Saccharomyces cerevisiae, yeast model for aging

In Yeast, Being Old Can Be a Good Thing

March 13, 2017

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A square slab can make an excellent door stop. Over time, though, the corners can get chipped, making the slab a bit rounded. This new bit of rock makes a less useful doorstop, but a much better wheel! The chipping and aging of the original stone has made it worse in some situations, but better in others.

A new study by Frenk and coworkers in Aging Cell shows that something similar can happen in yeast. Young yeast are much better at utilizing glucose, but older yeast have them beat with galactose (as well as with raffinose and acetate).

One way to think about this is that age turns yeast from a glucose specialist into a sugar generalist. Aging chips away at a yeast cell’s ability to use glucose, but this loss results in a gain in its ability to use galactose.

So at least in the right environment (i.e., when there’s lots of galactose around), with our old friend Saccharomyces cerevisiae, there can be advantages to getting older.

What makes this particularly fascinating is that at least in yeast, this suggests that there may be a positive selection for aging because of the advantage it can give in certain environments. Those yeast who are ageless would compete less well compared to their aging counterparts when their glucose was taken away. The aging process wins out over immortality!

Frenk and coworkers used a relatively simple experimental set up. Take young cells and old cells, mix them together, and see which outcompetes the other using various sugars.

They used yeast that had been aged for 6, 24, and 48 hours in glucose. This is a nice range as 6-hour “old” yeast are fully viable, 24-hour “old” yeast are starting to suffer a bit in the reproductive viability department, and the 48-hour “old” yeast have passed the median lifetime of a yeast cell. Young adult, middle aged, and elderly yeast.

In the first experiment, they compared these yeast to log-phase yeast which the authors refer to as young (vs. the other three which are referred to as aged).

While the 6 hour yeast could hold its own against the young yeast in glucose, the 24-hour and 48-hour yeast grew much more slowly. This is what you would expect, the younger yeast growing faster than the older yeast. The young guns outdoing the older generations.

The situation was different in galactose. Here, the elderly, 48-hour yeast, ran circles around the young yeast. They blew them out of the water.

And it appears to be an age thing. When they compared 6- and 48-hour yeast that were aged in galactose instead of glucose, the more aged yeast still won. So, it wasn’t the shift in environment that caused the difference, it was in the older yeast cells all along.

The change is also not permanent. The offspring of the older yeast weren’t any better at growing in galactose than the younger yeast were. Only the cells that had lived a longer life could use galactose so well.

A concern here is that yeast as old as 48 hours are pretty rare in the wild. But when they changed assays and looked at colony size as opposed to competition, they saw that even 18-hour yeast had an advantage over the young whippersnappers.

This was such a surprising result that they also looked at cell cycle times of individual aged cells and their daughters. The older mother cells cycled faster in galactose than their daughters. And the opposite was true in glucose.

So it really looks like there are advantages to growing older. Things break down a bit, but that breakdown uncovers new talents that had previously lain dormant.

If you’re a yeast, growing old is not a one-way decline into dotage. You gain new abilities that, under the right conditions, let you outcompete your children! The older cells are selected for in the right environments. #APOYG shows us something good about growing older.

by Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics

Categories: Research Spotlight

Tags: aging, evolution of aging, generalist, specialist

Life Needs to be More Like a 1950’s Chevy

October 21, 2015

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In the old days, a car came with the bare minimum of features to get from point A to point B. The windows rolled down with a crank and it usually had a radio. That was about it.

As the car has evolved, it has gained a huge number of bells and whistles. There are power windows and power brakes, a baffling number of computer-based bonus features, personal wifi hotspots, and so on. All of these have undoubtedly made cars more fun and comfortable to drive. But they have come at a cost. Many cars simply do not last as long as their predecessors because these extras break easily.

Turns out life may be like a modern car. It has lots of nice features that help it to do better in the world. But a lot of these features may shorten its life span.

This point was reinforced in a recent study by McCormick and coworkers. They painstakingly searched through a library of 4,698 single gene deletion strains in S. cerevisiae and found that 238 of these strains were able to produce significantly more buds over their lifetime. Many nonessential genes seem to shorten a yeast’s life.

And boy was it painstaking! Believe it or not, they manually dissected over 2.2 million individual yeast daughter cells to generate these results. Luckily it was worth it, as they found so many interesting things.

First off, many of the genes they found fall into a set of five pathways that includes cytosolic and mitochondrial translation, the SAGA complex, protein mannosylation, the TCA cycle, and proteasomal activity. So there are certain pathways we can target to extend the lifespan of our friend yeast. And even better, yeast may not be the only beneficiary of these studies.

Two of the pathways, cytosolic and mitochondrial translation and the TCA cycle, have also been found to be significant in extending the life of the roundworm C. elegans. These pathways are also shared with humans.

And just because the authors found no overlap with the other three pathways in other beasts doesn’t mean they may not be targets for life extension in them too. It could be that previous screens in C. elegans simply missed genes from these pathways.

It could also be that what is found in yeast may turn out to be important in people but not in C. elegans. For example, the authors failed to find any equivalent to the SAGA complex in C. elegans. Either the roundworm lost this complex during evolution, or the homologs between yeast and C. elegans are so different that they’re unrecognizable. In any event, humans at least do have an equivalent to SAGA, called STAGA.

All of this suggests that there may be common ways to make organisms, including people, live longer, healthier lives. Here’s hoping!

And these five pathways are certainly not the whole story. The majority of the genes McCormick and coworkers identified were not in these five, which means there are probably lots of other ways to get at living longer.

One fascinating example that the authors decided to look at in depth was LOS1. Deleting it had one of the biggest effects on a yeast’s reproductive life span.

At first this seems a little weird, as Los1p exports tRNAs out of the nucleus. As expected, deleting LOS1 led to a buildup in tRNAs in the nucleus. The authors confirmed that this buildup is important by showing that overexpressing MTR10, a gene involved in transporting tRNAs from the cytoplasm to the nucleus, led to a longer lived yeast with a buildup of tRNAs in its nucleus.

The next step was to figure out why having a lot of tRNA in the nucleus makes yeast live longer. It was known previously that Los1p is kept out of the nucleus under glucose starvation conditions. The authors confirmed this result.

Most everyone knows that restricting calorie intake (also called dietary restriction or DR) can extend the lives of most every beast tested so far, including yeast. The authors found that growing a los1 deletion strain at low glucose did not increase the lifespan of this strain any further. It thus appears that an important consequence of DR is keeping Los1p out of the nucleus and thereby increasing the amount of tRNA in the nucleus.

While we don’t know yet exactly why keeping tRNAs in the nucleus helps yeast live longer, it is interesting that the increased lifespan associated with the loss of LOS1 is linked to caloric restriction. Finding a way to inhibit Los1p has to be better than starving yourself!

This study has identified 238 genes to follow up on for future studies. And of course there is a whole class of genes that haven’t yet been investigated—the essential genes! Many of these may be important for extending life too. 

Stripping life down to its bare essentials may help individuals live longer at the expense of being the most fit in terms of survival in the hurly burly world of nature. After all, those “nonessential” genes undoubtedly have a function in helping yeast outcompete their less well-endowed yeast neighbors. Just like those power sliding doors are way better than the manual ones on a minivan.

But if you want a long-lived minivan, get the one with the manual doors. And if you want a long-lived yeast (or person), get rid of some of those nonessential genes that cause you to break down.

by D. Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics

Categories: Research Spotlight

Tags: aging, lifespan, Saccharomyces cerevisiae

Sharing the Health

April 22, 2015

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A study published a few years ago made a big splash in the health news by showing that obesity is socially contagious. If one person gains weight, their friends tend to gain weight too—even if they don’t live in the same town! This works the opposite way too: thinner people are more likely to be socially connected with thinner people.

You might think this is because people tend to make friends with others of a similar size, but this doesn’t seem to be the case. The researchers concluded that there is actually a cause-and-effect relationship: we all influence the weight of our friends.

Well, S. cerevisiae cells are not so different. They may not have social lives, but since they can’t move on their own, they do tend to live together in colonies. And within these colonies, they influence each other: not in terms of weight, but in terms of the effect that calorie intake has on the length of their lives.

Turns out that like nematodes, fruit flies and even mice, living on a meager diet makes yeast live longer. And in a new study published in PLOS Biology, Mei and Brenner found that yeast cells actually share the life-extending benefits of calorie restriction with their neighbors, probably via a still-unidentified small molecule.  

Yeast are normally grown in the lab on medium containing 2% glucose. To a yeast cell, this is like an all-you-can eat buffet that goes on for its entire lifetime. Media with a glucose content of 0.5% or less represent a meager diet. But that deprivation comes with a benefit, in the form of an extended lifespan.

Mei and Brenner already had some hints from previous studies that yeast cells might excrete a substance that promoted lifespan extension. To study this systematically, they devised an experiment to test whether mother cells change the media surrounding them as they divide.

The researchers placed individual mother cells in specific spots on Petri plates containing an all-you-can-eat buffet (2% glucose), a restrictive diet (0.5% glucose), or a near-starvation diet (0.2% glucose). They watched as the cells budded, and removed each new daughter cell as it separated from the mother, counting the buds. The lifespan of a mother yeast cell, termed the replicative lifespan, is measured as the number of times she can bud during her lifetime.

After the mother cells had budded 15 times, half of them were physically moved to fresh parts of the same plate, while the other half were left in place. For the mothers on the 2% glucose plates where calories were abundant, the move didn’t change anything. The mothers that were moved had exactly the same replicative lifespan as those that stayed put.

On the plates where calories were restricted, it was a different story. The cells that stayed in place had extended lifespans, as expected under these low-calorie conditions. But the cells that were moved to new locations lost most or all of the life extension—even though calories were still restricted in their new locations. This suggested that the mother cells had secreted a “longevity factor” into the medium surrounding them, which then extended their lifespan when they got older.

There were a couple of metabolites that were prime candidates for the longevity factor: nicotinic acid (NA) and nicotinamide riboside (NR). NA and NR are precursors to nicotinamide adenine dinucleotide (NAD⁺), a compound that acts as an essential cofactor for many important enzymes. They had already been implicated in lifespan extension because mutating genes involved in their metabolism can affect how long various creatures live.

When the scientists tried supplementing calorie-restricted cells that had been moved to fresh medium with either NA or NR, they found that supplying these metabolites could restore the longevity benefit.  This finding strengthens the idea that NAD⁺ metabolism is involved.

But was the longevity factor actually NA or NR? To test this, Mei and Brenner grew yeast in liquid media with the different glucose concentrations and then tested for NA and NR in the medium using liquid chromatography-mass spectrometry analysis.  They found that under all the conditions, the amount of NA secreted by the cells didn’t change and secreted NR was undetectable, suggesting that neither was the factor induced by calorie restriction.

To ask directly whether there is a diffusible longevity factor, the researchers grew cells in liquid medium containing 2% or 0.2% glucose until all the glucose was used up, then separated out the cells and freeze-dried the remaining liquid. They suspended the dried “conditioned” medium in water and spread it on plates to repeat the cell-moving assay.

Just like before, cells grown in 2% glucose had the same lifespan after being moved to a fresh spot, and the addition of resuspended conditioned medium to the plate didn’t change that. However, the starved cells grown on 0.2% glucose not only kept their lifespan extension when moved to conditioned media, but actually lived 10% longer compared to starved cells on un-conditioned media that were not moved.

When the researchers dialyzed the conditioned medium so that molecules smaller than 3.5 kDa were lost, the longevity factor was lost too. So it looks to be a small molecule, and of course they are actively pursuing its identity. Intriguingly, this would explain why other scientists have been unable to detect calorie restriction-induced lifespan extension in yeast using microfluidic technology, where immobilized yeast cells are grown with a constant exchange of growth medium. Under these conditions, a small molecule that promotes longevity would be washed away.

So, even though they don’t have Facebook friends, yeast cells influence the health of their peers. Rather than spreading the influence through social interactions as we humans do, they broadcast a chemical that is the key to long life. 

It’s tempting to think that the identity of this chemical will tell us something about human aging. But if this mysterious molecule worked in humans the same way as it does in yeast, people would still have to eat just enough food to stay alive to get the benefits. Still, perhaps the molecule can point us towards finding a treatment that will let us live longer while enjoying lots of good food. We could have our cake and eat it too!

by Maria Costanzo, Ph.D., Senior Biocuration Scientist, SGD

Categories: Research Spotlight

Tags: aging, calorie restriction, NAD+, Saccharomyces cerevisiae

Not too long, not too short

October 02, 2014

Have you ever noticed that the length of your hair is just right for a small window of time? Too short, and you feel a little exposed – too long, and it gets in your face.

Like your hair, the telomeres at the ends of your chromosomes have a length that is just right for them (and you). Telomeres are non-coding sequences of DNA added to the ends of chromosomes that protect the important DNA there from being lost. Telomeres that are too long may contribute to cancer; while telomeres that are too short are associated with aging.

It makes sense, then, that telomere length should be carefully regulated (just as, in a perfect world, you might want to keep your hair the perfect length all the time). The enzyme that adds the non-coding DNA sequences to the end of chromosomes, telomerase, is composed of highly conserved subunits. In yeast, this enzyme is a quaternary complex composed of the regulatory subunits Est1 and Est3, the catalytic subunit Est2, and the RNA template component, TLC1.

Previously, it was thought that telomerase was primed for action whenever it was needed in the cell. But this does not seem to be the case.

In a recent publication in Genes & Development, Tucey et al. showed that, in addition to the active quaternary telomerase complex, two subcomplexes – a preassembly complex and a disassembled complex – were also present in the cell. Each of these subcomplexes lacked one subunit compared with the active telomerase, and the missing subunit was different for each subcomplex.

So, how did the authors discover these subcomplexes? Using a special immunoprecipitation protocol, one of the first things the researchers noticed was that Est1, Est2, and the TLC1 RNA were associated with each other throughout G1 and S phase, but that Est3 was missing from the complex during G1 phase and present only at very low levels throughout S phase. They called this Est1-TLC1-Est2 complex the “preassembly complex.”

In fact, Est3 was only appreciably associated with the preassembly complex during G2/M and late in the cell cycle. And, even then, only 25% of the preassembly complex was associated with Est3 to form the active quaternary complex. So the presence of the active holoenzyme was regulated with respect to the cell cycle.

Since Est3 was not always bound to the preassembly complex, the authors set out to determine what was required for Est3 binding. They found that neither Est1 nor Est2 could bind Est3 if it was unable to bind TLC1 RNA. Thus, the RNA component of the presassembly complex was necessary for its two other subunits to form an active telomerase by binding to Est3.

In addition, they determined that both Est1 and Est2 contained binding surfaces for Est3, and that both of these surfaces were required for the full association of Est3 with the preassembly complex. So Est3 interacts directly with each of the other protein subunits to form the active telomerase.

There was still one more question that intrigued Tucey and coworkers – Est3 appeared to exist in excess compared with the other subunits, so why did it have such limited association with the rest of the telomerase subunits? This suggested some sort of inhibitory regulation of Est3.

Indeed, the authors uncovered an Est3 mutant, Est3-S113Y, which appeared to have lost the ability to be inhibited. This mutant exhibited elongated telomeres and increased association with telomerase, and was associated with the preassembly complex in G1 phase rather than being restricted to late in the cell cycle. This mutation lies directly adjacent to Est3’s binding domain, leading the authors to conclude that Est3 is regulated by a “toggle switch” that specifies whether or not it can bind to the preassembly complex.

As mentioned previously, the authors also saw evidence of a second subcomplex during their studies. When they drilled down on the components, they identified a “disassembly complex” that lacks only Est2, in contrast to the preassembly complex that lacks Est3. They determined that this subcomplex is inactive and requires the prior formation of the quaternary complex since its formation requires Est2 binding to TLC1, just as is observed for the preassembly complex.

Given the cell’s desire to have telomeres that aren’t too long and aren’t too short, it makes sense that the enzyme that lengthens them is regulated. There is a toggle switch that signals formation of the active complex and, once formed, the active complex is transient, dissociating its catalytic subunit to become inactive. This regulation ensures that telomerase is active only late in the cell cycle. Just as you might work to keep your hair just the right length, the cell regulates telomerase to keep the lengths of telomeres from getting out of control.

by Selina Dwight, Ph.D., Senior Biocurator, SGD

Categories: Research Spotlight

Tags: aging, protein complex, telomerase, telomere

Yeast, the New Fountain of Youth

January 23, 2014

Ponce de Leon searched the New World for the fountain of youth.  Turns out that if he had some of the tools at our disposal, he wouldn’t have even had to leave Europe.  He just needed to go to the local bakery or brewery and look inside the yeast he found there.  Of course, then he wouldn’t have found Florida…

Using in silico genome-scale metabolic models (GSMMs) in yeast, Yizhak and coworkers identified GRE3 and ADH2 as two genes that significantly increased the lifespan of yeast when knocked out.  Even more importantly, their method also allowed them to identify the mechanism behind this increased lifespan—the mild stress of increased reactive oxygen species (ROS).  This last finding may help scientists identify drug targets that they can target to increase the lifespan of people too.  If only Ponce de Leon had lots of -omics data and a powerful computer or two!

After constructing an in silico starting state, Yizhak and coworkers entered two sets of data from previous work that had been done on aging in yeast.  They next used gene expression profiling to identify which metabolic reactions were different and which were the same in young and old yeast.  They then systematically tested the effect of knocking out these reactions one at a time in their computer model to identify those that could potentially transform yeast from old to young with minimal side effects. 

Their first finding was that many of their best hits, like HXK2, TGL3, and FCY2, had already been identified as important in prolonging a yeast cell’s life.  They decided to look at seven genes that had not been previously identified as being involved in aging. 

When two of these seven, GRE3 and ADH2, were knocked out, these yeast strains lived significantly longer with minimal side effects.  For example, the strain lacking GRE3 lived ~100% longer than the wild type strain.

Figuring out why these yeast probably lived longer was made simpler because they used metabolic models to identify the genes.  The hormesis model of aging suggests that mild stress, like that found in caloric restriction, can lead to increased life span.  With this model in mind, the authors focused in on the possibility that knocking out GRE3 and/or ADH2 would lead to increased stress through the production of increased levels of ROS.  When they looked, they found that the two knockout strains did indeed have higher levels of two common forms of ROS, hydrogen peroxide and superoxide. 

Of course none of us is particularly interested in extending the life of a yeast!  But these results could suggest new drug targets to go after that might mimic the effects of caloric restriction without us having to starve ourselves.  And these same methods can be used on human cells to find key pathways to target in people.  In fact, the authors have started to use their computer models to investigate aging in human muscle cells and found that like in yeast, many of the genes they have identified are consistent with previous work on human aging. 

Now we probably shouldn’t get too far ahead of ourselves here.  This is a promising first step but it really isn’t much more than Ponce de Leon boarding his ship to begin his trip to the New World.  We still have a long voyage ahead of us before we find the fabled fountain of youth.  

by D. Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics

Categories: Research Spotlight

Tags: aging, metabolic model, Saccharomyces cerevisiae

Oxidation: Maybe Not SO Bad After All

July 11, 2013

You don’t have to be a scientist to get the message that oxidation is bad and antioxidants are good. Just go to the vitamin aisle of your local supermarket, or listen to the ads on late-night TV.  You’ll quickly find out that oxidation caused by free radicals is the reason for aging, and antioxidants are the fountain of youth.  Of course you shouldn’t believe everything you hear…

Things just aren’t that clear when you take a good hard look at aging. Yes, oxidation happens, but there actually isn’t solid experimental proof that it causes aging. In mice, this connection has not panned out at all: lowering the ability to sop up oxidants, by knocking out an antioxidant enzyme, does not shorten the mouse’s life.

In a recent eLife paper, joint first authors Brandes and Tienson and their coworkers used our favorite experimental subject, Saccharomyces cerevisiae, to see if oxidation is a cause or just a consequence of aging. They generated a ton of data about oxidation during aging and did not find any evidence for causation.  Instead they came to the surprising conclusion that the trigger for aging may actually be a sudden drop in the levels of the coenzyme NADPH.

The first step, published previously by this group, was to come up with a very sensitive assay for protein oxidation. The amino acid cysteine can act as a sensor for levels of oxidation, as its sulfur-containing thiol group can be oxidized and reduced. Their technique, known as OxICAT, detects the ratio of reduced to oxidized thiol groups on cysteine residues for individual proteins. They can do this for hundreds of proteins at the same time.

In the current study, they looked at the oxidation state of cysteine residues in about 300 different proteins and also measured the levels of several different metabolites related to the redox state of the cell. All of these data were collected over time in aging yeast cells, both under normal conditions and under conditions simulating caloric restriction or starvation.  These last conditions were included because a lower-calorie diet has been shown to slow down aging, in yeast as well as in animals.

Oxidation of proteins definitely did increase over time. But if oxidation were the cause of cell death, you would expect that it would increase steadily and at some maximum point, the cells would die. Surprisingly, that didn’t happen.

Instead, different groups of proteins were oxidized with different kinetics. The most sensitive proteins (about 10% of the set that they studied) were oxidized 48 hours before the cells started to lose viability. This set included some conserved proteins that are important in maintaining oxidation-reduction balance in the cell, such as the thioredoxin reductase Trr1p. 

But it wasn’t only those especially sensitive proteins that were oxidized. In a second wave of oxidation, almost all the remaining proteins (80%) were oxidized at 24 hours before death. And even with so many proteins oxidized the cells were still metabolically active, with ATP levels near normal. So massive oxidation did not equal instant death for these cells.

As predicted, a low-calorie diet slowed down the whole process. The pattern looked a lot like it did in cells on a normal diet, but there was more time between the waves of oxidation and before the end of viability.

The authors also looked at what happened to different metabolites during aging. One key metabolite is the coenzyme NADPH: it donates electrons to the thioredoxin system that helps balance oxidation and reduction. They found that even before any changes in oxidation are detectable, levels of NADPH decrease very suddenly. The authors speculate that this decrease starts the collapse in redox potential that ends in the death of the cell. The oxidation of protein thiols is an effect rather than a cause, and could actually be a way for the cell to sense its redox state and possibly regulate it. NADPH levels have been seen to decrease in aging rats as well, suggesting that this could be a universal part of the aging process.

The results of this study are too voluminous to describe fully here, but they raise a lot of intriguing questions. Some proteins never got oxidized – what protected them? Are NADPH levels really the trigger for aging, and if so, what causes the sudden decrease? Is oxidation of cysteines actually part of a sensory mechanism? And if that’s true, would preventing oxidation really be such a good thing? This may be another good reason to turn off late-night TV.

by Maria Costanzo, Ph.D., Senior Biocurator, SGD

Categories: Research Spotlight

Tags: aging, oxidation, redox, Saccharomyces cerevisiae

Studying Aging in Yeast Just Got Easier

April 02, 2012

Watching a yeast cell age can be a real pain.  In budding yeast like Saccharomyces cerevisiae, the buds quickly outnumber the mom.  Which means scientists need to remove the buds as they appear.

Up until now, scientists have had to use a 50-year-old method that involves removing the buds by hand.  Not only is this labor intensive, but the field is held back by the inability to use high resolution microscopy to investigate the aging process. 

These technical limitations may soon be swept aside with a new microfluidic dissection technique described by Lee and coworkers in a recent study out in PNAS. These researchers were able to monitor 50 aging yeast at once with a variety of microscopic techniques without having to remove the buds by hand.  And unlike the older technique, they were able to keep a constant environment for the yeast cells (i.e. no decrease in nutrients and/or build up in wastes).

Basically Lee and coworkers tucked the yeast mother cells under a micropad which they then washed with a constant flow of nutrients.  Because the daughter cells are smaller than the mother, they are washed away as they emerge.  So no manual bud removal is required.

Sounds convenient but the researchers needed to show that this new technique gave similar results as compared to the old one.  And they did.

They showed that mutant strains behaved similarly with both techniques.  So a SIR2 deletion mutant still had a shorter lifespan and a FOB1 deletion mutant still lived longer with microfluidic dissection.  Not only that, but the number of divisions in an average yeast’s lifetime was comparable with both techniques.  At first blush the techniques do seem comparable.

Now they were ready to take their new technique out for a spin to see what it could do.  First they were able to show heterogeneity in how yeast cells age.  Some cells died as spheres around their 12th division while others died as ellipsoids after their 25th division.  The shape of the yeast later in life correlated with how long that yeast lived.

The researchers were also able to use GFP to explore the vacuoles of aging yeast.  They found three classes of vacuoles: tubular, fused, and fragmented.  The tubular vacuoles were only found in the longer-lived ellipsoid yeast.

Researchers could not have discovered these properties of aging yeast without the new microfluidic dissection technique.  And these findings are really just the tip of the iceberg of what can now be learned about aging by studying yeast.  It will be exciting to see what else scientists will be able to learn about the twilight of a yeast cell’s life.

Life and Death of a Single Yeast

by D. Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics

Categories: Research Spotlight

Tags: aging, microfluidic, vacuole, yeast

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