The stress of life

What is stress for you? Last-minute gift shopping? An upcoming presentation? Or maybe riding the underground during rush hour? 

We get stressed when the world presents us with a challenge and it is an experience we share with all other living beings. While we humans worry about matters big and small, the microscopic realm is the stage of a deadly arms race between microbes and viruses. Plants are also under constant pressure – they must fend off pathogens, parasites and predators while at the same time optimising their photosynthesis despite constant changes in temperature and light. How does life cope in an ever-changing environment? 

Using the example of microorganisms and plants, microbiologist Isabelle Zink as well as biochemist and systems biologist Wolfram Weckwerth from the Department of Functional and Evolutionary Ecology at the University of Vienna explain the amazing anti-stress strategies of life. Some of them also provide possible answers to questions that challenge our own adaptability – from medicine to food security and climate change.

Microbes under stress

What a bacterium goes through in its lifetime can be shown using the best-researched of all microbes: E. coli. It thrives at 37°C in the intestines of humans and other warm-blooded animals. Outside of this comfort zone, however, trouble looms. "From a biological point of view, stress is any change that disrupts the natural balance within a cell," explains microbiologist Isabelle Zink. "When exposed to heat, the proteins in the cell lose their specific shape and therefore their function. Cold conditions, in turn, often change the shape of the mRNA (messenger RNA), which is why it cannot be properly read and 'translated' into proteins."

Both are bad news for bacteria, since proteins are the molecular machines that control processes in the cell. "The cell then produces protective proteins," explains Zink. "These can be, for example, heat shock proteins that help other proteins to maintain the correct shape. On the other hand, there are also cold shock proteins that ensure that mRNA can be read despite the cold." This allows the cell to survive within a range of temperatures, even in adverse conditions. Heat above 60°C, however, sooner or later kills E. coli and most other bacteria, which is how pasteurisation of food works.

Stress is relative

Isabelle Zink's favourite single-celled organisms, however, are different. Archaea may look like bacteria, but from an evolutionary perspective they are more closely related to us humans. And some of them are true survival experts. 'Hyperthermophilic' ('heat-loving') organisms thrive in extremely hot environments, specialising in temperatures above 80°C. One species of archaea even reproduces at 122°C on the walls of deep sea 'underwater volcanoes'. Others can handle extremely acidic, alkaline or salty waters, high concentrations of heavy metals, great pressure or radioactivity. Thermococcus gammatolerans, also a deep-sea dweller, can survive radiation levels more than a thousand times higher than those lethal to humans.

Such examples show that what is extreme for some microbes is totally normal for others. On the other hand, thermophilic archaea are unable to reproduce at 37°C – the temperature which is perfect for E. coli. Stress is relative, depending on the respective ecological niche. 

Microbes vs. viruses and the discovery of genetic scissors

However, nowhere are bacteria and archaea safe from their biggest enemies: viruses. They are not living beings but instead tiny packages of infectious genetic material (DNA or RNA) found in every corner of the world. Isabelle Zink illustrates their numerical dominance. "Prokaryotes – i.e. bacteria and archaea – are the most common cells of all; there are about nine times as many of them as there are stars in the galaxy. But viruses even outnumber them, as there are on average up to 13 viruses for every microbe." 

Microbes have also developed immune systems to keep viruses at bay. One of these is CRISPR-Cas, which is often referred to as 'genetic scissors'. As this name suggests, CRISPR is a molecular machine that can cut DNA. By cutting, for example, the DNA of a virus, it renders this virus harmless. 

Although microbes have many virus defence systems (see info box), CRISPR-Cas is special, explains Isabelle Zink. "It 'remembers' a virus species that has already attacked the cell before and fights the intruder even more fiercely if it returns – similar to the way people form antibodies after vaccination. CRISPR is like a vaccination certificate for microbes." 

Tools for medical diagnostics

Isabelle Zink has dedicated her work life to the defence systems of archaea. She is currently intrigued by the possibility that CRISPR could not only defend against viruses but also control how strongly genes are expressed in the cell. "There is also the question of how different defence systems interact," adds Zink. "Each microbial species has on average at least six different defence systems. So far, we have only looked at the systems individually. But we need to look at the immune system as a collective – that is where I want to take my research."

The microbial 'firewalls' are also interesting for diagnostic applications. During her Schrödinger Fellowship project in Wageningen in the Netherlands, Zink patented an 'Argonaut defence system' for this purpose. "If you want to identify a specific gene variant in a sample, such as a cancer-causing mutation, our tool makes it possible to detect it even in the tiniest amounts. The Argonaut system specifically degrades the DNA of the healthy gene variants in the test tube while at the same time maintaining the mutation. This means that the mutation can be recognised better, and perhaps earlier, in diagnostics." Forensics, where gene variants are used to identify individuals, is another possible field of application.

Plants: Green survival artists

The easiest way for our E.coli bacterium to avoid stress is to swim away. Plants, in contrast, are sessile and stay in one place for their entire lives – but that is precisely what makes them so interesting, says biochemist and systems biologist Wolfram Weckwerth: "Due to their sedentary nature, plants have developed some of the most complex cellular adaptation processes. This explains their success: plants make up at least 80% of the Earth's total biomass, which is approximately 450 billion tons of carbon. Bacteria and archaea make up approximately 80 billion tons of biomass, and animals, including humans, about 2 billion tons."

The example of drought stress shows just how complex the strategies of the green survival artists are. Hot conditions or a lack of water cause stressed cells to produce more of the plant hormone abscisic acid. This initiates processes that prevent further water loss. The microscopic stomata in the leaves close, through which plants normally inhale the carbon dioxide required for photosynthesis but also lose water through evaporation. The hormone also stimulates the growth of roots and their ability to transport water, while at the same time inhibiting shoot growth. 

The formation of the hormone is, in turn, controlled by the interplay of countless signalling molecules. "This involves what are known as protein kinases – enzymes that trigger a cascade of cellular signals resulting in a stress response," explains Weckwerth, a trained chemist. "These are also active when the plant's immune system defends against viruses, bacteria or fungi. On average, plants have three to four times as many protein kinases as humans. This shows how diverse the molecular mechanisms are with which they react to stress."

Pearl millet: A grain fit for the future

We humans are well advised to better understand the stress biology of plants, emphasises Weckwerth, namely "in order to ensure regional and global food security, and thus individual and planetary health." Globally, our diet is chiefly based on a small number of crops such as wheat, rice, maize and soya, also indirectly as animal feed. This means an unbalanced supply of calories and nutrients. Secondly, in order to grow, plants extract CO2 from the air through photosynthesis. This makes them the largest carbon sink on Earth and thus a global factor in limiting global warming." Crucially, the efficiency of photosynthesis depends on the plant's stress resilience.

As the world's population is set to grow until the end of the century, the farming industry of the future will have to feed more people on less land. "We therefore need to stabilise and increase yields – despite climate change, which is increasingly stressing our plants," explains the biologist, who is a member of the research networks 'Environmental and Climate Research Hub' (ECH) as well as 'Health in Society'. 

Common varieties of crops such as wheat have been bred for high yields but are also susceptible to stress. Weckwerth and his team are therefore interested in what are referred to as 'orphan crops'. These are varieties which have great future potential but are currently used only locally. One of these is pearl millet, which is particularly drought-resistant and is grown in places including the Sahel region. It also has a much higher iron content compared to common types of grain.

In field trials the researchers have tested whether pearl millet could also thrive in Austria. "The seeds had problems with the cold spells in spring," explains Weckwerth. "Then we sowed in May and – lo and behold – the millet germinated, grew faster than maize and we were able to harvest at the end of August." 

The search for climate-resistant crops

So what makes pearl millet so special? In dry conditions, its photosynthesis is more efficient than that of native grasses such as wheat. This is partly due to the special anatomy of its leaves and partly to its ability to prevent what is known as 'chlorophyll degradation'. While a lack of water causes wheat leaves to wither, pearl millet remains green. Weckwerth and his team were able to break down the profile of the active signalling pathways and proteins in pearl millet that are characteristic of this 'stay-green effect'.

"In the long term, we want to use the fundamental knowledge we are currently developing to enable breeding programmes that improve food security," says Weckwerth. "We have decoded several genetic signalling pathways that mediate drought resistance. With the help of our partner organisations, who have the world's largest collections of varieties, we can benefit from the genetic diversity of existing varieties. For example, we can identify wheat varieties that have the required resistance signalling pathways. These traits can then be cross-bred into the common wheat lines to increase their stress resistance."

Plants and their microbiome: A superorganism

The omnipresent microbes also play a role in another aspect of plant health which, to this day, has received little attention in research. The inside and outside of a plant and the soil around its roots are home to bacteria, archaea and other microorganisms. "In fact, the evolution and physiology of plants are so closely linked to their microbiomes that we can consider it a 'superorganism', also known as a 'holobiont'," explains Wolfram Weckwerth. 

The soil microbiome appears to play a decisive role in plant performance. On the one hand, there are microbes that help the plant to grow. Through their metabolism and signalling molecules they indirectly promote plant growth or provide resources for plants such as phosphate or nitrogen. On the other hand, some microbes remove nitrogen from plants by converting ammonium to nitrate, which is quickly washed out of the soil into the groundwater – a massive environmental problem in modern farming.

Plants, on the other hand, appear to give a huge amount of resources to their soil microbiome. "Through their roots they release large volumes of photosynthetically produced substances into the soil," says Weckwerth. Our studies indicate that plants 'control' their microbiome via these metabolites, known as root exudates – the substances attract beneficial microbes and suppress the growth of undesirable ones.

Optimised plants for stressed ecosystems

"The ability to control the substances excreted through their roots appears to give plants a way to improve their resilience," says Weckwerth. His vision is to improve this ability in cultivated crops. As with drought resistance, the researchers are looking at the genetic diversity of existing varieties. "In a panel of 50 different wheat lines, we were able to demonstrate an enormous variability of root exudates. Each line also had its own individual microbiome."

In a large-scale study with 300 different wheat lines, the researchers and their international collaborators now want to find out which plant genes and signalling pathways control the soil microbiome. This knowledge can be used as part of breeding programmes to develop resilient varieties with an optimised microbiome. 

"These varieties can reduce microorganisms that take nitrogen away from them, known as nitrifiers, because they produce inhibitory root exudates," Weckwerth cites as an example. This gives these plants a better nitrogen uptake capacity and at the same time means that less nitrate enters the groundwater. Such crops in the fields would be an asset for higher yields and in the fight against the environmental crisis caused by overfertilisation. At the same time, beneficial microbes must not be suppressed, such as those that convert methane, a very powerful greenhouse gas, into less harmful CO2.

The complexity of plant-microbe relationships shows that the more we know about these two very different but closely intertwined life forms, the more we can benefit from their amazing adaptations. "We must not forget one thing," Wolfram Weckwerth emphasises, "the plant kingdom binds about 157 billion tons of CO2 every year, including man-made CO2. If we want to get a better grip on climate change, we need to work with this ecosystem."