Annually, an estimated 1 billion people suffer from critical fungal infections, with 1.6 million deaths. The impact of fungal diseases, however, is much greater. 16% of crops lost globally result from microbial diseases, with up to 80% of these caused by fungi. Additionally, agricultural animals are severely affected, resulting in low yields and early deaths. With the release of Oxfam’s Hunger report over the summer, highlighting that by the end of the year 12,000 people could die of hunger per day as a result of COVID-19, the impact of fungal outbreaks on agriculture cannot be underestimated.
Drug resistance in fungal infections is becoming increasingly challenging as there are few effective treatments available. There is a widespread use of antifungal agents in agriculture, promoting the selection of resistant mutants. This highlights how vital it is to understand how fungi acquire drug resistance.
A research group at the Wellcome Centre for Biology in Edinburgh, led by Professor Robin Allshire, discovered that the fission yeast Schizosaccharomyces pombe can acquire resistance through epigenetic changes, i.e. changes to the activation state and regulation of DNA rather than the DNA itself. This was demonstrated by using genome sequencing, which identified no genetic changes in the genes the researchers were investigating.
The group used caffeine to create a hostile environment, mimicking the effects of antifungal drugs, and showed that S. pombe cells can adapt to adverse conditions without genetic changes. Instead, resistance is acquired through the formation of heterochromatin, i.e. regions of silenced genes due to the addition of chemical modifications to DNA organising proteins (histones). This prevents proteins necessary to activate genes from accessing the DNA, resulting in gene silencing. As these mutations are epigenetic, the resulting drug resistant mutants are referred to as epimutants.
We chose to use caffeine as an insult because caffeine resistance is conferred by the deletion of genes with a variety of cellular roles, thus, increasing the chance of obtaining epimutations.Allshire et al., 2020
The newly formed heterochromatin reduces the expression of key genes which ultimately results in caffeine resistance. Forcefully inducing heterochromatin in these genes confirmed that heterochromatin formation leads to this resistance.
The epimutants created are unstable mutants, as the histone modifications are reversible through the actions of an enzyme (Epi1). In the absence of Epi1, the heterochromatin can be inherited by daughter cells after cell division, forming stable mutants, in which the mutation cannot be reversed. When unstable epimutants were grown in the absence of caffeine for 14 days, 23% of cells lost caffeine resistance, indicating unstable resistance can indeed be reversed.
Previously, researchers searched for drug resistant mutants using genome sequencing which would exclude mutants that acquired resistance epigenetically, as they show no changes in their genome sequence. Professor Robin Allshire’s group, however, selected for weak mutants which had not yet acquired genetic changes, thereby investigating a previously unknown mechanism of developing drug resistance.
Importantly, epimutants showing unstable caffeine resistance were also found to be resistant to some antifungal agents, including clotrimazole, tebuconazole, and fluconazole, all of which are widely used antifungal agents. This suggests that resistance to antifungal mechanisms can arise by the same mechanism. These new insights could be the first step in the development of new antifungal therapies for resistant infections. The authors suggest repurposing drugs to target histone modifying enzymes needed for heterochromatin formation, or pursuing the development of drugs targeting fungal, but not host, heterochromatin formation.
Re-engineering existing so-called ‘epigenetic drugs’—compounds that inhibit histone-modifying enzymes—or searching for novel agents of this type may identify molecules that specifically block the formation of fungal, but not host, heterochromatin, reducing the emergence of antifungal resistance in clinical and agricultural settings.Allshire et al., 2020
These findings promise to accelerate the development of new, more efficient drugs for resistant fungal pathogens, targeting the epigenetic changes directly. This provides hope of keeping up to speed with the evolutionary arms race between drug resistant pathogens and drug discovery, and thereby alleviating fungal diseases as well as increasing crop yields.
Written by Giovanna Weykopf and Tara Wagner-Gamble, and edited by Ailie McWhinnie.
Giovanna’s thoughts… Epigenetics has been my passion for years, and the relevance of epigenetics is only increasing with many new discoveries being made in this area. This finding by Allshire et al. revolutionises how we investigate drug resistance. It opens many new promising approaches to drug development which is especially important in the context of rising drug resistance and the number of crops lost annually to fungal pathogens. This study clearly shows that epigenetics has implications in many different areas previously unsuspected. It will be exciting to see a clearer picture of the wider importance and mechanisms of epigenetic regulations emerge within the coming years.
Find Giovanna on LinkedIn @Giovanna Weykopf
Tara’s thoughts… The integrated relationship between hosts, pathogens, and their genomes in the context of drug resistance is vital to understand given the rise in drug-resistance seen across all types of infectious diseases. This study clearly highlights our emerging understanding of the role epigenetics plays in acquired resistance and will hopefully lead to new classes of drugs which inhibit or reverse unwanted epigenetic changes. Fungi are a challenge to treat anyway, due to their similarities to other eukaryotic cells, therefore, we don’t want to lose the effective drugs we have. This is an exciting area of research, and I can’t wait to see where it leads.
Find Tara on LinkedIn @Tara Wagner-Gamble