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Tristan Free
Unraveling some of the mysteries of fungi with a novel gene-editing tool, researchers have identified a number of molecules with promising anti-cancer properties.
Scientists from the University of Pennsylvania (PA, USA) have introduced fPE7max, a prime editing platform optimized for fungi, which they’ve used to isolate a library of complex molecules, many of which possess new-to-science structures. Among them, are three with possible tumor-busting prowess, presenting an exciting opportunity for future drug discovery efforts.
Despite their renowned contributions to modern medicine, fungi have historically been overlooked, especially in genetic research. This is in large part due to the fact that they switch off their innate antimicrobial gene pathways when grown in the lab.
“To turn those silent pathways back on, we needed a powerful way to precisely manipulate fungal genomes, such as editing their master regulatory genes, but traditional tools weren’t up to the task,” study author Xue Gao explained.
CRISPR, for example, is not optimal in filamentous fungi – major ecological contributors and industrial hosts characterized by tubular filaments known as hyphae – as the dominant DNA repair pathway in these organisms is non-homologous end joining, which rapidly stitches broken ends together and is inherently mutagenic, rather than the more accurate and methodical homologous recombination. As a result, the double-strand breaks introduced by CRISPR are more likely to lead to unintended mutations. Prime editing offers an alternative, capable of targeted gene editing without double-strand breaks. However, prime editing had never been established in filamentous fungi before – until fPE7max came into the picture.
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The novel tool integrates fLa, a stabilizing protein, to protect guide RNA from degradation that can accompany large DNA insertions and deletions, as well as a protein that bypasses the fungus’s natural repair system to ensure that edits are not reversed.
The resulting platform supports a range of different edits, from single-base substitutions to kilobase-scale insertions and multi-kilobase deletions (up to 10 kb), with an average editing efficiency of almost 90% across diverse genes and species.
Putting fPE7max to the test, the team targeted translation-regulatory elements upstream of the metabolic regulator gene laeA. Metabolomic profiling revealed activation of previously underexplored biosynthetic pathways, which led to the identification of 18 new fungal metabolites. Eight of these, the researchers believe, have structures that had never been reported before, and three demonstrated cytotoxic activity, making them potential candidates for further exploration in cancer drug discovery.
“These molecules can serve as lead compounds for disease treatment, providing a vital new pipeline for drug discovery,” first author Chunxiao Sun asserted, adding that one of the new molecules showed selective toxicity against human breast, hepatic and leukemia cancer cells.
“It’s a compelling proof-of-concept demonstrating that the next generation of life-saving therapeutics might already exist in nature,” Gao added.
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