There is simply a certain quantity of stress biological structures can withstand before they crumble. Tissue fluids freeze into ice crystals when exposed to temperatures below −3 °C, for example, and enzymes break down and grow to be dysfunctional at extremely low temperatures. But there are organisms which can be built to thrive in such harsh conditions. Some species of fungi, for example, can survive the cruel weather in Antarctica, and scientists have spent years attempting to work out how. A study published in Science Advances on September 7 suggests that these polar organisms can have adapted because of tweaks to unstructured regions of their proteins.
The intrinsically disordered regions (IDRs) of proteins are the formless, liquidlike parts of proteins that lack the power to fold right into a functional shape and sometimes react with RNA to form naked, or membraneless, cell organelles akin to the nucleolus through a phenomenon referred to as liquid-liquid phase separation (LLPS).
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The study shows that yeasts adapted to harsh cold experienced evolutionary changes in IDRs, which change how phase separation occurs. The researchers found that the structure of the IDRs in species adapted to polar regions is different from those in temperate areas.
The researchers came upon these differences while studying how transcription occurs within the cold. They were analyzing two primary components of the transcription system of RNA polymerase II multisubunit enzyme—carboxy-terminal domain (an IDR) and Ess1 prolyl isomerase—in five cold- and salt-adapted species of yeast isolated from the Arctic and Antarctica, once they noticed that the species’ carboxy-terminal domain’s (CTD’s) structure was barely different than that of the model yeast species baker’s yeast (Saccharomyces cerevisiae).
Steve Clabuesch, a former colleague of study coauthor Steven Hanes, walks along Commonwealth Glacier, McMurdo Dry Valleys, Antarctica in 2016.
In baker’s yeast, the CTD consists of a repeating peptide sequence—YSPTSPS—whereas the polar yeasts’ repeating sequences diverge at positions one, 4, and 7. Study coauthor Steven Hanes, a molecular geneticist at SUNY Upstate Medical University in Latest York, says that the repeating sequence is shared across vast clades of life and is sort of similar even in humans, so this divergence in cold-adapted yeasts was “extremely significant.” He and his colleagues were interested by why the polar yeasts’ CTDs manifest such differences and wondered if these CTDs would still function in baker’s yeast.
First, Hanes and colleagues deleted the gene that codes the CTD of baker’s yeast but kept the CTD intact by retaining the plasmid that expresses the enzyme Rpb1 (a subunit of RNA polymerase II), which kept the host cell alive. Next, they cloned a gene for polar yeasts’ CTD right into a plasmid and transferred it into baker’s yeasts. They did this to check whether the divergent CTD would work when paired with the structured region of RNA polymerase II in baker’s yeast. The procedure was carried out at 18 °C and 30 °C to find out the consequences of colder temperatures.
Hanes explains that if the cloned CTD gene are compatible with the host, they’ll push out the unique plasmid from the host’s cell in favor of the brand new one. The degree to which the originals are lost will give an estimate of the cloned plasmid’s degree of compatibility with the model species.
The baker’s yeast replaced its own plasmid with that of the varied polar yeasts fairly well at 30 °C. But at 18 °C, the baker’s yeast containing CTDs from the Arctic fungi Wallemia ichthyophaga, Aureobasidium pullulans, and Hortaea werneckii lost only 0.2 percent, 13.6 percent, and 21.5 percent, respectively, while people who contained CTDs of the Antarctic fungi Dioszegia cryoxerica and Naganishia vishniacii didn’t lose any of the unique plasmids. In contrast, the control lost 58 percent and 87 percent of the unique plasmid at 18 °C and 30 °C, respectively. The researchers report that the CTD from polar yeasts didn’t work in baker’s yeast at 18 °C.
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Hanes says that he suspected the differences may stem from the mechanisms of LLPS, as a previous study revealed CTDs can undergo the method. So the team checked if the CTDs from the cold-adapted yeast may undergo this phenomenon.
The researchers sure polar CTDs with purified proteins in a test tube and checked for LLPS by observing the cloudiness of the answer—evidence that LLPS occurred—at different temperatures and salinity levels. Hanes and his team observed that the CTDs of those polar yeasts do undergo phase separation, but they accomplish that in another way than baker’s yeast. They noticed that the CTDs of species that were most compatible with S. cerevisiae at 18 °C exhibited high LLPS, while people who weren’t compatible showed none. The researchers attributed these differing properties to the divergence within the CTDs’ amino acid sequence, they usually hypothesize that this divergence might promote cold and salt tolerance within the polar species.
Hanes says that the intrinsically disordered regions within the protein with more variable sequences are very adaptive to “selective pressures that may change the biophysical properties of how the proteins sort out inside cells.” And this adaptation can change how, when, and where they undergo phase separation.
“We all know that stress can induce phase separation by certain proteins, but what we’re suggesting is that environmental tuning of phase separation allows organisms to tolerate temperature and other extreme conditions,” says Hanes.
Amy Gladfelter, a cell biologist on the University of North Carolina who studies phase separation and didn’t work on the brand new study, says that the outcomes are “really suggesting that by natural variation and . . . at how free-living yeast may adapt to extreme temperatures and likewise extreme salinity, [the research team] can find evidence of adaptation in sequences which can be essential to drive phase separation.”
Hanes tells The Scientist that the study has uncovered more questions than answers, however the team intends to resolve most of them, including the precise mechanism through which phase separation properties confer environmental tolerance, because this might help other microorganisms to survive harsh and changing climate conditions.