The Beal and Went experiments are steeped in history, but they are not merely exercises in scientific nostalgia. Even after all this time, they are still generating valuable data in pursuit of a question that remains unanswered: how long do seeds live? Perhaps there are as many answers to this question as there are seeds, but even rough trends by genera, family or species remain elusive. There is still a lot we don’t know about how long seeds last.
This is a problem because no matter how well they are stored, all seeds eventually die. One study even demonstrated that seeds age, albeit very slowly, when stored in liquid nitrogen, which has a temperature around −196°C. It seems that, even at temperatures more than two-thirds of the way to absolute zero, things will steadily fall apart. It also seems that the rate of ageing is highly variable between the seeds of different plant species, something observed in the Beal and Went experiments so far, as well as numerous other studies. Furthermore, even within species, different varieties can have quite different life spans. Back in 2005, Christina Walters and her colleagues at the USDA analysed the viability of numerous samples held in the genebank there, including some that had been collected as far back as 1934. A few plant families tended to have long-lived species with scarcely any change in seed viability after decades. A couple of other families had more than their fair share of short-lived species, with most seeds dead after a similar time period. But, for the most part, longevity didn’t seem to fall along family lines. The researchers also noticed that some seed life spans clustered together depending on the seeds’ original habitat, with several Australian species lasting longer than some European species. But it was difficult to find reliably predictive patterns. Over the years, every time a general rule almost manifests, along comes a bunch of confounding exceptions.
To this day, the longevity of any one seed from any one species can be hard to predict. It is largely through observational studies that examine the life spans of seeds on a species-by-species basis that the picture is gradually being filled in. For example, the seeds of lettuce, peanut, willow or elm do not live as long as the seeds of wheat, mung-beans or chickpeas. Given the same storage conditions, poppy seeds will outlive cabbage seeds, and white lupine seeds will last longer than celery seeds, and you’re doing well if you get even a few years out of chive seeds. Sorghum and barley seeds will certainly outlive rye seeds, and peas will likely outlast them all. It’s tempting to say, ‘Well, that’s genetic diversity for you,’ but it’s even more complicated than that.
Katherine Whitehouse is a seed physiology specialist at the Australian Grains Genebank who studies longevity in orthodox seeds. She tells me that life span can vary significantly among seeds from the very same species collected in the same location in different years. ‘We could have two seed lots that you germinate before storage and they’re both 98 per cent germination,’ she says. ‘They’re the same species and the same variety, but they were grown, for example, in different years. They both went in [storage] at the same time, so you would assume they were the same.’ But that’s not necessarily the case. ‘One will lose viability three times faster than the other one,’ says Whitehouse.
Scientists at seed genebanks routinely test the viability of their holdings by removing a subset of the seeds from a particular accession and attempting to germinate them. The percentage of seeds that germinate provides a glimpse into how well the rest of the seeds in that sample are doing. It’s sort of like polling. They do this test just before the seeds are put into storage, and then they test at regular intervals thereafter. Obviously, this needs to be stretched out – every five years, every ten or even every twenty if things are still going well, otherwise the entire accession can be used up with testing. ‘We do regular monitoring,’ says Mariana Yazbek at ICARDA. ‘Every five years you open that package, you take fifty seeds and put them in a petri-dish, and you germinate them and you count how many of them are alive.’
Although the life span of any given seed is not easy to predict, and general rules for species or families are often elusive, it is now clear that all orthodox seeds follow a remarkably similar pattern as they age. ‘Imagine you drew a graph where the X-axis was storage time and the Y-axis was viability,’ says Christina Walters. She lifts one hand and begins to trace an invisible horizontal line in the air, just above her eye-line. ‘The real difficulty of working with seeds and ageing is that everything is fine. It’s fine,’ she continues, following that line, which is still fairly flat. ‘It’s fine. It’s fine. It’s fine, and then, all of a sudden, it crashes.’ Her hand drops as she traces a precipitous slope, one that tapers off at the bottom as if she’d just been describing the path of a steep ski run. ‘There’s this threshold and a crash,’ she reiterates. ‘But that time period before it goes bad, you have no symptoms.’
And there’s the rub: most seeds look the same whether they are alive or dead. Unless they’re very obviously damaged or hollowed out by deterioration or disease, you just can’t tell. You remove a seed after three years of storage, or after thirty years, and you hold it in the palm of your hand. Maybe it’s dormant, maybe it’s kind of middle-aged, maybe it’s No Longer With Us. ‘You can’t tell!’ exclaims Walters. ‘That’s why I got into them. It’s like they have a history but they’re not going to reveal it.’
Seed ageing is a major focus of global genebanks, says Charlotte Lusty at Crop Trust. ‘You’re monitoring the collection because you need to be on top of the viability of all of those seeds and all of those packets,’ she says. ‘If at any time viability starts to go down, then there’s a danger of it going down very quickly. At the moment, the threshold that is used is 85 per cent. So if less than 85 per cent of the seeds in the packet are proving to be non-viable … it could be that very quickly, they all become dead.’
That’s when you need to act fast, says Yazbek. ‘You don’t want to get to that tipping point where you’re suddenly conserving dead seeds … So we’re monitoring very carefully to make sure they’re still living or not, and once that tipping point is reached, we put [the seeds] out in the field, we plant them and we harvest fresh seeds.’
It’s not a perfect system, but at present it’s the most effective way to keep genetic lines going, says Lusty. Nevertheless, it entails a lot of work and resources. ‘It means that if you have a very large collection, you need to have a big team of staff who are periodically checking each seed packet,’ Lusty says, adding that it’s a ‘massive data exercise’ in which everything must be catalogued and tracked precisely. ‘Inevitably,’ she says, ‘when you have an old collection, it means every year you have to plant out a percentage of [it] … in order to create new fresh stock that can go back into storage for another few decades. Plus, you’re also dealing with hundreds of requests for seeds, so they’re coming out and you’re sending seeds off, doing all the paperwork. So, while you can imagine it being a relatively simple job of keeping seeds in jars, it actually ends up being a very intensive business when you’re doing it on a large scale.’
Lusty also worries about genetic erosion within individual accessions because even just one packet of seeds can contain a lot of interesting genetic variety. Certain crops are self-pollinating, so the seeds stored in any one accession tend to be somewhat similar genetically, she explains, but there are a lot of crops that don’t fit this profile. Maize, for example, is cross-pollinated, so genes are contributed by different parent plants. Lusty clarifies that she’s not talking about modern-day sweet corn, the kind you buy in the grocery store and which has been carefully bred to be ‘totally uniform’. She means the old maize landraces. ‘In an ear of maize of a traditional landrace, all the little seeds are different colours just in one ear’, and you can literally see the different traits, the genetic diversity right before your eyes, she tells me. It’s like an entire genetic population all growing on the same plant. If you put all those seeds just from that one ear into a seedbank accession, ‘it’s like a completely diverse set of individuals that make up that accession. There’s as much diversity in that one accession [as] there is between the accessions.’ When even some of the seeds in that accession start dying, you’ve lost that unique arrangement of genes forever.
Seed life spans are an issue for all genebanks, including the biggest seed storage backup of them all: the Svalbard Global Seed Vault. Despite how eternal that arctic fortress appears, nothing in there will last forever. Yet, once the seeds arrive, they are never tested. ‘Genebanks are recommended to ship high quality seeds only,’ says Svalbard’s Seed Vault coordinator, Åsmund Asdal. Svalbard is a mirror site, after all, so the genebanks that deposit the seeds must regularly monitor their primary collections. ‘They have the same seeds from the same yield, same quality at home,’ he says. ‘So they test these seeds and when they see that these seeds are getting old, and do not germinate well enough, they know that they also have to replace the seeds in the Seed Vault.’
The intervals for testing can vary. It might be every ten years or every twenty-five years, says Asdal. That might sound like a nice breather between tests, but there are so many seeds and so many starting points for storage. Saving lots of seeds from different years is a good thing, as it provides what genebankers like to call ‘temporal diversity’, but the downside is that seedbanks are perpetually testing their seeds. Will the barley seeds in a particular accession last 400 years or will they just scrape past twenty? What about the same variety collected a decade earlier during a drought? And what of the barley wild relative from Azerbaijan? Knowing the answers to questions like these would save a lot of time and resources that instead could be invested in saving more seeds of more species.
For this reason, scientists working with the Svalbard Global Seed Vault want to better understand seed longevity. And in a similar vein to the Beal and Went experiments, they’re running their own long-term studies. ‘In 1986 we started a so-called 100-year project aimed at investigating [the] longevity of seeds stored in permafrost,’ Asdal tells me, explaining that they produced and packed a new set of seeds for this experiment and placed them in Coal Mine No. 3, not far from the original seed collection. ‘The purpose of that has been to monitor how long can these varieties stay alive in permafrost at −3°C.’
While it is interesting to see how long the seeds of different crops stay alive and germinate in permafrost conditions, Asdal cautions that it doesn’t reveal very much about the longevity of the seeds in the vault because the coalmine conditions are too different. ‘Because seed longevity is of vital interest for the genebanks and for us,’ he says, ‘we have decided to introduce a new 100-year seed longevity experiment in the Seed Vault, where the temperature is −18°C.’ The experiment focuses on fourteen crop species that are important to global agriculture, including wheat, barley, peas and lettuce – and for each crop species, five varieties are included. The samples are being provided by six genebanks in varying locations to ensure geographical diversity. The first seeds were deposited at the Svalbard Global Seed Vault in August 2020, and Asdal tells me more seeds will be produced and deposited over the next three to four years, ‘to cover the significance of different growing conditions’. In other words, temporal diversity.
Intriguingly, one of the biggest threats to seed longevity is the same metabolic reactions that are necessary for life. During cellular metabolism, genes are activated and cellular components are built and broken down and built again. Energy is consumed, converted, stored and released. This happens in all functioning cells. It’s just how they traverse existence. It’s how all cells live. But it’s also how they die. Not so long ago cell metabolism and cell death were believed to be independent phenomena: one provided building blocks and energy, the other just kind of tore it all down, and each was controlled by a largely different sets of genes and molecules. But it turns out they are tightly linked. For example, many proteins known to play central roles in metabolism are now also implicated in cell death. These days, cell metabolism and cell death are referred to as ‘deeply intertwined’, with their pathways comprising a ‘tangled circuitry’.
It makes sense, really, that the biochemical processes of life and death can’t be extricated from one another, and that there is nothing so clear-cut as a section of the genome for life and another for death. It’s still not entirely clear what flips the final switch, only that many of the same components are involved. So, where there is metabolism, everything required for death is already in play, ever ready to pivot towards that final endeavour.
Normal metabolic activity – including metabolism – involves a lot of cellular motion. Cells are filled with a kind of gelatinous fluid called cytoplasm, where organelles and biomolecules move around, shifting and diffusing this way and that. There is a lot of matter in there, but also enough molecular elbow-room for efficient movement so that it behaves as a fluid system. There are collisions, reactions, molecular messages swiftly being sent and received, genes being transcribed into proteins, proteins slipping past one another as they move this way and that, and cell structures being built, repaired, broken down, recycled or expunged. In short, most cells are crazy busy, and actively metabolising plant cells are no exception.
During all this activity, chemical bonds are formed and broken, leading to the production of free radicals, which are atoms or molecules with an unpaired electron. It seems such a tiny problem – it’s literally subatomic – but anything with an unpaired electron is a proverbial loose cannon. Most atoms, except hydrogen, far prefer their electrons to come in pairs. ‘Prefer’ is probably the wrong word here, though. It’s more of a demand for which there is zero negotiation. Fundamental quantum forces are at play and they won’t take no for an answer. As such, an atom with an unpaired electron is an unstable little beast and downright ravenous for another electron to make a pair again. It will do anything to get one, and often that means thievery.
Enter oxygen, much lauded for the vital role it plays in the production of cellular energy and many other important biochemical interactions. But it turns out that an oxygen molecule (O2) is easily split into two unstable oxygen atoms, each with an unpaired electron. So, in biological systems, where a lot of oxygen is being shuffled about, most of the free radicals are reactive oxygen species (ROS). As they careen all over the cell, they slam into other molecules and, in a process called oxidation, the ROS steals an electron, ripping it away from where it was happily residing. The ROS, now sated, is no longer reactive, but now the other molecule is missing an electron and the problem continues. During metabolism, vital biomolecules such as proteins, lipids and DNA are bombarded with ROS. Electron by electron, these free radicals chip away at these important biomolecules, and in so doing they gradually erode a cell’s structure, function and genetic stability.
Cells do their best to mitigate all this oxidative damage. They use enzymes such as superoxide dismutase, which essentially eats free radicals for lunch. They also activate genomic repair mechanisms, marshalling troops of specialised enzymes that detect damaged DNA and quickly fix it. Cells will use other enzymes to produce small molecules called antioxidants. And this is a neat trick. An antioxidant molecule is able to donate an electron, yet it will not become a voracious electron scavenger itself. Vitamin E, a well-known antioxidant, achieves this by donating a hydrogen atom, which only has one electron. The idea is to have enough antioxidants floating around that they’re more likely to be hit than anything vital – they kind of take one for the team.
In all living cells, there is a constant battle between free radicals and the cellular machinery tasked with either preventing or mopping up the damage. Cells cannot keep this up forever, though, and there comes a point where damage outpaces repair. Eventually, cells die. As long as cellular metabolism is in full force, the biological clock is ticking steadily, relentlessly.
Plants cannot slow down time itself, but in orthodox seeds they have found a way to profoundly slow the effects of metabolism under certain environmental conditions. From the seed’s point of view, this amounts to almost the same thing. During seed development, there comes the seed filling stage. This is when the seed accumulates all that dry matter it’s going to need – lipids, carbohydrates, proteins, fats and so on. At this point, the seed still has a vascular connection to the mother plant, and as such, there is still fluid available. But as more and more dry matter accumulates, the ratio of dry matter to fluid begins to change. Things are still happening in a fluid state – genes are active, cellular metabolism is chugging along – but it’s all happening in an increasingly more crowded space.
When the connection between an orthodox seed and the mother plant is cut a new phase begins. It’s called ‘maturation drying’ and a lot changes inside the seed. First of all, the water in the seed evaporates until it reaches an equilibrium with the humidity in the air, so if the relative humidity in the surrounding environment is low, the seeds lose a lot of moisture. ‘The seed is going to dry down and the further removal of water compresses all of those cellular constituents,’ says Christina Walters. All that once-busy cytoplasm now looks very different, she says. ‘It’s just kind of a mishmash of material compressed so far that it holds its shape.’ The cellular contents become so crowded that the flow of molecules inside the cells slows down. Molecules that would zip past each other in a fluid are now compressed together and – this is key – they begin to interact. Katherine Whitehouse explains that this ‘leads to really strong bonds between them.’ These bonds further reduce mobility of the molecules. And, at low enough moisture levels, the result is a state that very closely resembles glass.
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Look around you right now and there’s a good chance you’ll find yourself in the presence of some glass. Whatever the item – a drinking glass, glass lenses, a window pane – it was once a molten liquid that reached temperatures of over 1700°C and was then allowed to cool. During this process, the molecules never achieved a structured arrangement but instead remained as disordered as they had been in the liquid state. A molecular snapshot of the liquid state and the glass state would appear almost identical, although there is more crowding in the glass thanks to the contraction that occurs during cooling. As this happens, molecules have a harder time slipping quickly past one another, so they begin to slow down, and as they do this, they make stronger connections. It’s a bit like how it’s easier to grab the hand of someone who is walking slowly past you than to grab the hand of someone driving past you in a convertible going 150 kilometres per hour, which is not advisable. As the cooling continues, so, too, does this process until at last the molecules become fixed in place and glass is formed.
There is an old ink bottle on a windowsill in my home which was made sometime in the mid-1800s and is long since empty of ink. The thick, greenish glass catches the morning light and is full of lovely imperfections frozen in time. When I hold it up, I can see tiny bubbles that have been trapped in place since the day the glass cooled. Inside the bottle, the bottom is thicker on one side than the other, as if the piece had been carelessly left to cool at a slight angle and now there is a tiny slope of glass caught forever in the act of rising. No matter how much I tip the bottle the other way, it will not even out in my lifetime, but that’s not to say it never would. This highlights an important notion: the glass state is non-equilibrium. In other words, glass is neither a true liquid nor an ordered crystalline solid, but rather occupies a place somewhere in between. Exactly where in between is still a matter of debate. Glass is classed as an amorphous solid to distinguish it from a crystal, but physicists have been arguing about the true nature of glass for years, and if you were inclined to start an argument in a bar full of materials engineers, then just ask loudly whether glass is more of a solid or a liquid. Recently, materials engineers Edgar Zanotto and John Mauro proposed a new definition of glass that accounts for profoundly gradual changes. A glass, they say, might seem solid but it’s actually very, very slowly relaxing towards the liquid state. The critical element is time.
The point is that plants that produce orthodox seeds figured out how to use glass physics to their advantage. ‘I think they’re the original materials engineers,’ says Christina Walters. Orthodox seeds, with their accumulation of food stores and subsequent drying out, achieve a ‘glassy’ state, she explains. Like the molecules in glass, the molecules in seed cells slow down and become trapped in an amorphous, disordered state. Like traditional glasses, seeds are not in equilibrium: they are no longer liquid, but they are not crystalline solids. And for the seed, this is a good thing. Being fluid would make extreme life spans impossible, yet being in an ordered crystalline state would be disastrous. Among other things, the resulting solid would be far too stable. The entire point of a seed, after all, is a sort of useful impermanence. A seed is meant to carry genes from one point in time to another, and then its job is done. By being neither liquid nor fully solid, a seed has a chance to change. ‘It’s not totally fixed. That’s the beauty of it!’ says Walters.
Inside glassy seeds, motion is mostly small-scale and short-range. Glucose is no longer barrelling around at high speed and proteins have slowed to a halt, yet molecules can still react with their neighbours ever so slightly. There are also random pores throughout this glassy solid, through which gases and water molecules can slowly diffuse. ‘You might think that as you increase the proximity of reactants, then you would increase the reaction rate,’ says Walters, ‘but because it’s so solid and packed and the molecular mobility is so slow, you’re slowing those reactions.’ And that’s the whole idea, she tells me: ‘The reason for solidification is to slow down reactions.’
Enzymes are central to metabolism, but they can only function if they can reach the molecules they’re meant to interact with, says Walters. ‘The chances of catalysis through an enzyme are pretty small because how are you going to bring those molecules together?’ And so, metabolism is suspended and the seed becomes quiescent. This is how, under the right conditions, an orthodox seed slows down the effects of time.
Ultimately, a seed’s longevity is a trait, one acquired early in its life. The health of the mother plant and the environmental conditions during seed development can have a big impact on longevity, says Katherine Whitehouse. She explains that it is crucial that the seed experiences neither too much stress nor too little during this time, because there’s a sweet spot that triggers a series of important stress responses that will profoundly influence how long that seed can live. It is an elaborate interplay between genes and the environment. As for exactly which genes in which species, which types of environments, and precisely when and how they interact, well, that’s something that a lot of plant physiologists are keen to find out.
It appears that just after the connection with the mother plant is cut and the maturation drying phase begins, the seed prepares a lot of proteins and other molecules it will need to endure the drying-down phase, as well as a potentially long period of quiescence. For example, many seeds produce proteins called late embryogenisis abundant proteins (LEAs), which are kind of weird. They’re unstructured proteins like slightly unravelled string, but they seem to act as a kind of buffer against dessication, helping to keep other molecules in the seed stable in dry conditions. Some seeds also produce heat-shock proteins that help protect other vital proteins from temperature-stress. This is also a time when the seeds ramp up their supply of antioxidants.
For seeds that enter a glassy, fully quiescent state after drying out, it’s not clear how long they can remain that way in a soil seedbank. After all, periods of rain come and go, daily humidity rises and falls, dew gathers, fogs may drift in, perhaps a light snow falls and melts. As Whitehouse explains, ‘the moisture content of the seeds in the soil seedbank is constantly changing, fluctuating with the environmental conditions.’ This alters the seeds’ physiology quite a lot, she says. Water relaxes a seed out of the glassy state and the amount of water present can influence which reactions can take place or the degree to which metabolism returns.
It is interesting to note that humans are not the only ones who make seedbanks. Plants do it, too. Out in nature a seed-bearing plant will drop seeds onto the soil, creating what is known in ecology as a ‘soil seedbank’. The seeds remain there until a particular suite of conditions arrive, and provided the seeds are still viable at this point, they will germinate.
Louise Colville is a seed biologist at the Millennium Seed Bank at Kew Gardens in the United Kingdom, and is particularly interested in the longevity of the seeds of wild plant species. She tells me that changing moisture levels can actually be quite helpful to a seed. ‘You get cycles of rehydration and dehydration, and during rehydration phases the damage to DNA or proteins can be repaired. That is one aspect of why seeds can actually survive for quite a long time in a soil seedbank.’
There are a lot of intriguing questions about where seed dormancy comes into all this. The physiological choreography involved in cycling in and out of dormancy and keeping track of environmental cues would appear to be impossible in a completely dry, glassy seed. Just how dormancy is broken in a seed that’s in a glass-like, quiescent state is still being worked out, and it has been called ‘one of the great mysteries of plant science’.
In a 2019 paper in Frontiers in Plant Science, Leeds University plant scientist Wanda Waterworth and her colleagues proposed that dormancy and glassy quiescence may actually be two distinct strategies for long-term survival and that, over the course of evolution, plants in different environmental niches might have gone with the one that worked best. They reason that seeds from wetter environments can carry out more cellular repair, but because there’s more rainfall or humidity those seeds are more likely to need dormancy to help control the timing of germination. Meanwhile, seeds from drier environments wouldn’t need as much help in that department, but they would need a lot more protection from long-term damage.
For seeds enduring long periods of dry conditions, the dry, glassy state is a useful defense mechanism. It’s likely that the glassy state is a significant factor in many cases of extreme longevity. Elaine Solowey sprouted the 2000-year-old date palm seeds from Masada and the caves near the Dead Sea, just at the edge of the arid Jordan Desert, certainly a place where there are long droughts. One of the more recent droughts, she tells me, lasted twenty years. ‘We had 20 millimetres of rain, which is like spitting on the ground twice,’ she says with a laugh. Whenever she has tried to germinate ancient date palm seeds from other locations, nothing ever sprouts. ‘No seed that I have ever planted that was ancient came up unless it was from the Dead Sea area,’ she says. ‘I think there’s something special about that area. I’ve tried old seeds from other places, and as far as I’m concerned they were dead as doornails!’
By drying seeds down and storing them at low temperatures, seed-banks take advantage of orthodox seeds’ defense mechanism by coaxing them into a glassy state and then keep them there. How then, do seeds age even when carefully stored at low temperatures and low humidity? Because the glassy state comes with a few downsides, says Christina Walters. ‘You are bringing molecules [together] that are reactive, like a sugar molecule has an O-H group that definitely wants to give up an electron. It’s going to give that electron away and something is going to receive it. So you’re still going to be making free radicals.’ These free radicals have a lot more trouble moving around, but the smaller ones, such as single activated oxygen atoms, can still squeeze through the jam-packed glassy matrix like pickpockets on a crowded subway. As they degrade other molecules, new free radicals can be formed. This all happens very slowly now, but it still happens.
Then there’s just the nature of the glass itself. In much the same way that the strength of any glass can range from fragile to strong, research by Walters and others has shown that strength of glassy-state in seeds can vary. It seems that, among other things, the types and amounts of proteins and other molecules that are packed inside the seed influences how well, or how poorly, the glass forms. That certainly leaves a lot of room for differences between species or the influence of environmental conditions while the glassy solid is forming. Even within any one seed, there are all sorts of differences in glass strength precisely because it’s so disordered and messy in there. It’s also full of imperfections, like tiny gaps in a crowd. ‘What you have produced is this matrix with lots and lots of pores, and nothing likes a vacuum, so you’re always going to have that compression of the pores,’ says Walters. As the glass ages, the bonds that locked all the molecules in place gradually relax and the small-scale structure of everything shifts a little and the pores are compressed. Molecules that had been separated become close enough to react. So, new free radicals are produced slowly but inevitably. This is how seeds age, says Katherine Whitehouse. Over time, these free radicals do what free radicals always do, she explains. They destabilise membranes and degrade genetic integrity.
Ironically, as long as the seed remains in a glassy state, it cannot repair itself because the enzymes that would otherwise detect and carry out such repairs can’t move. To protect against deterioration, seeds make antioxidants prior to entering the glassy state and these bear the brunt of free-radical oxidation. That’s why so many seeds are rich in antioxidants – they’re even extremely useful in seeds that never become glassy, like recalcitrant seeds. But so long as a seed is in a glassy state it can’t make new antioxidants, and if enough time passes, they will run out.
For Walters, the intriguing mechanisms of seed ageing have been ‘a lifelong fascination’, and there is still a great deal that remains unknown. She and others have spent years conducting chemical post-mortems on dead seeds, looking for the key event that pushes a seed down that precipitous slope that finally causes it to die, a sort of molecular coup de grâce. They have wondered if the critical tipping point is the destruction of the mitochondria, or the depletion of all the food reserves, or maybe there is some important molecule that finally fails. They’ve been looking for the analyte that will give it away. An analyte, explains Walters, is an identifiable chemical that can be detected, and which, in this case, might point to a specific cause of death. If there was a single common cause of death in seeds, ‘you would expect something very specific to go down as you see things die, or go up as you see things die. We have a lot of dead things and we never find the same analyte!’ says Walters, adding that in test after test, in regards to the chemical read-outs: ‘What you usually see is a mess!’
But Walters realised that this mess was telling them something important: ‘If you think about it, the seed, during embryo development, accumulated all this dry mass. Every single molecule in that cell is now subject to deterioration, and that’s a lot of deterioration. There’s lots and lots of different ways it can deteriorate because different molecules have been pressed together. So, where I’ve evolved to is this is exactly what we should expect: a mess!’
It evokes an intriguing analogy with human ageing. There’s this idea that a person can die of ‘old age’, says Walters. ‘But what is that? Usually someone falls and it goes bad, or something else bad happens. That brought us to this idea that lots of little events have tiny, tiny effects, but then you have a big effect from a small event, a chance event.’ In other words, she explains, ‘Many, many things can go wrong as you age, but that’s not the lethal event. The lethal event could be a minor event in another circumstance, but because there are so many things going wrong, [it] has a big effect. It’s the straw that breaks the camel’s back – in itself, it’s insignificant, but it could have a large consequence in that moment. It’s kind of humbling to think that’s how you live.’
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In order to store seeds long term, to prevent genomes from eroding and species from flickering out of existence, genebank operators have to anticipate the moment that seeds in an accession will begin to die and refresh them before that tipping point is reached. ‘All we have at the moment to estimate longevity or quality of seeds are germination tests,’ says Katherine Whitehouse, which, she concedes, is ‘a very crude measure’. A germination test doesn’t tell you what a seed’s longevity is, she points out. ‘It tells you the viability of a seed lot at that time but it won’t tell you how long it will stay at that germination [rate] before it will start declining, and that’s the problem.’
Moreover, some seeds might not germinate, not because they are dead but because they are in a state of deep dormancy. This isn’t really an issue for most crop species because they lost the dormancy trait during domestication, says Whitehouse, but many wild species and crop wild relatives still retain that dormancy trait. And if this isn’t broken by the right conditions – bushfire smoke, or maybe a journey through an orangutan’s gut – then it’s unlikely to germinate no matter how much water and light are offered.
In recent years, Christina Walters and her colleagues have been looking for something that can provide more information about ageing and longevity in seed collections. They have searched for molecules that will change over time as a seed ages, she says. They wanted something that could be sequenced because modern technologies would make analysis simple. DNA wouldn’t do because it’s structurally quite strong, so it wouldn’t provide the strong signals of change that they were looking for. They thought about searching for proteins, which can also be sequenced, but that kind of analysis requires large amount of proteins – or, as Walters puts it, ‘buckets of material’ – which you wouldn’t be able to get from tiny seeds. Ultimately, they settled on RNA.
RNA is an essential part of cell growth and metabolism. Its main job is to carry genetic instructions from the cell’s nucleus to another part of the cell where the instructions are carried out and all the cell’s vital proteins are made. As a seed develops, it makes heaps of RNA in anticipation of all the proteins and molecular machinery it’s going to need later when it’s time to repair damage, break dormancy or trigger germination. But in orthodox seeds that dry down to a glassy state, everything slows to a halt before all those genetic instructions can be converted into proteins by the ribosomes, which are the cell’s protein-making factories. In the glassy state, when the ribosomes stop working – everything does. This is an important strategy on the part of the seed, because with so much RNA already processed and ready to go, the seed can get to work really quickly when moisture levels rise.
Walters tells me that when the seed enters the glassy state, all that RNA becomes fixed in place just like everything else. It just so happens that RNA isn’t anywhere near as stable as DNA; it’s quite fragile. For one thing, DNA is double stranded whereas RNA is only single stranded, making it a lot more vulnerable to attack from radicals. Another clue is in the names: RNA stands for ribonucleic acid, while DNA stands for deoxyribonucleic acid. Both contain an almost identical core sugar molecule, but the ‘deoxy’ in DNA signifies that its sugar is missing an oxygen atom. RNA, by contrast, still has this oxygen atom, which is in an O-H group. That O-H group is pretty reactive, which helps RNA carry out its myriad functions in the cell. The catch is that it can easily react with another part of the RNA, causing it to break. The bottom line is that RNA is incredibly prone to oxidative attack and other causes of breakage – much more so than DNA. In a happily metabolising cell, that’s not such a huge deal because cells will get rid of damaged RNA and quickly make more. The ephemeral nature of RNA affords all living cells a certain agility in responding to environmental cues. But in the glassy state of the seed, that can’t happen. Any RNA a seed has before it enters that state is what it’s stuck with. As RNA gradually breaks down, due to oxidative attack and other damage, the cell accumulates broken fragments of RNA, like shards of broken pottery that cannot be swept away. Walters and her colleagues realised that they could measure the decline in RNA quality to see how quickly a seed is ageing. Moreover, it’s easy to isolate, measure and sequence, and you don’t need much of it at all, says Walters. ‘It’s such a nice analytical tool.’
Walters tells me that she has been analysing RNA quality in seeds to see how it changes over time. The quality of the RNA is signified with an RNA integrity number, or RIN for short. It’s a 10-point scale, and anything with high-quality RNA, such as fresh seeds, score at least an 8. But if the RNA breaks down, this RIN value drops. By the time it reaches 3, the RNA situation is pretty messy.
Walters wanted to know how RIN values change as seeds age, for which she needed a whole bunch of seeds representing a variety of different ages, from new ones to old dead ones. Walters tells me she didn’t have to wait around for seeds to die because she’s been collecting seeds for her research ever since she first started at the USDA. ‘Now, thirty years later, I have a lot of dead seeds!’ she says with a laugh. Recently, she and her colleagues selected soybean seeds spanning an age range from one to twenty-seven years, and sure enough, their RIN values declined with age and correlated well with decreasing seed viability.
This research is providing a window into what’s going on inside ageing seeds. For seedbanks, it promises to be a useful way of detecting the degree of ageing in seeds that have no external indicators of decline, which in turn will be valuable for managing long-term seed storage and, by extension, the preservation of crop and wild plant species. It’s also proving rather useful in outer space.
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On 2 September 2015, a Russian Soyuz spacecraft was launched from Kazakhstan carrying three humans, equipment, supplies, and precisely 2 kilograms of seeds. What species, you ask? Eruca sativa, better known as rocket. The seeds were part of a European Space Agency research project called Rocket Science. Get it? Don’t ever say cosmonauts and space botanists won’t seize any opportunity for a good pun. The seeds were kept on the International Space Station (ISS) for six months, then returned to Earth where, with the help of around 600,000 schoolchildren, they were tested for germination and the growth of any seedlings was monitored. Meanwhile, scientists at Royal Holloway, University of London, led a series of more detailed studies – from gene transcription analysis to RNA analysis – to see whether six months off-planet had caused accelerated ageing in the seeds.
There was a good reason for doing all this. Crewed space travel requires food supplies, and when space travel begins to involve very long distances, such as journeying to Mars or beyond, replenishing those food supplies will pose a major challenge. Seeds can facilitate space exploration by making it possible for crew members to grow their own food on long-haul space flights. Moreover, the transportation of seeds to Mars offers an opportunity to not only survive a trip to the red planet, but also to initiate Martian agriculture on arrival. Closer to home, it might show that space gardens could be useful for long-term habitation on the moon. However, none of this will eventuate if seeds themselves can’t survive long periods in space. For this reason, space agencies are keen to find out how long seeds live in space, how well they germinate, and whether the resulting plants have any difficulties in growing and producing more offspring. There are already a lot of unknowns surrounding the physiology of seed longevity on Earth, and spaceflight merely adds a whole new bunch of confounding variables, from the potential effects of microgravity and the mechanical vibrations during rocket launch, to the effects of ionising radiation from solar winds and galactic cosmic rays which exist beyond Earth’s magnetic cocoon at intensities no seed – nor human, for that matter – ever evolved to handle.
The findings of the Rocket Science project suggested that the rocket seeds on the ISS had subtly aged faster than the Earth-bound control seeds during their six-month journey. The researchers concluded that it might have something to do with the fact that the seeds on board the ISS had absorbed around 100 times more radiation than the seeds that had remained on Earth. ISS orbits Earth, high in the magnetosphere, so it still receives a lot of protection from our planet’s magnetic field, but a spacecraft journeying to Mars would be much more vulnerable – the ionising radiation exposure would be around five-fold greater. The study’s findings indicate more protective storage conditions will be needed, but still, it seems that one day seeds will be able to boldly go where no seeds have gone before.
And so, seeds, and our ability to preserve them, offer a promising link to a future in which we can reach other worlds, while protecting the biodiversity and food security on this one. They also offer us a precious connection to our past.