Killer plasmids as drivers of genetic code changes during yeast evolution

Project Information

CODEKILLER

Grant agreement ID: 789341

DOI

10.3030/789341

Project closed

EC signature date 25 May 2018

Start date 1 October 2018

End date 31 March 2025

Funded under

• EXCELLENT SCIENCE - European Research Council (ERC)

Total cost

€ 2 368 356,00

EU contribution

€ 2 368 356,00

Coordinated by

UNIVERSITY COLLEGE DUBLIN, NATIONAL UNIVERSITY OF IRELAND, DUBLIN
Ireland

Periodic Reporting for period 4 - CODEKILLER (Killer plasmids as drivers of genetic code changes during yeast evolution)

Reporting period: 2023-04-01 to 2025-03-31

Summary of the context and overall objectives of the project

The aim of the CODEKILLER project was to investigate a possible mechanism by which the genetic code can change during evolution. The genetic code is the set of rules that determine how the 64 different codons in mRNA are translated into the 20 different amino acids in protein molecules, for example the codon AUG is translated into methionine. In the cell, the genetic code is physically implemented by tRNA molecules, which are covalently attached to an amino acid at one end and make basepairs with codons in mRNA at the other end. The genetic code is often considered to be universal because it is the same in the nuclear genomes of most species, but in 2018 we discovered that the genetic code has changed several times during the evolution of budding yeast species. Specifically, the codon CUG, which is translated as leucine in most species, is translated as serine or alanine in some groups of yeasts. Because this change is almost the only evolutionary change in the genetic code that has ever happened during the evolution of nuclear genomes, our goal was to understand how it happened. Our hypothesis was that the evolutionary reassignment of the codon CUG was caused by natural selection imposed by a killer toxin, specifically a ribonuclease toxin that cleaves and destroys tRNA-Leu molecules with the anticodon CAG, which translate CUG codons as leucine. We postulated that many different killer toxins that cleave specific tRNAs may exist in yeasts, even though such toxins are rare and only 4 of them were known at the outset of the CODEKILLER project.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

By a combination of data-mining, targeted genome sequencing, and advanced bioinformatics methods, we increased the number of known toxins from 4 to more than 100. We found that killer toxins of this type are much older and present in a much broader range of fungal species than previously known. We found them in filamentous fungi and “early diverging” (basal) fungi as well as budding yeasts. We found that toxin genes can be located either in the nuclear genome, or on small virus-like elements in the cell’s cytoplasm.

We showed that many of the new toxins we discovered are functional, and we identified the tRNA targets of new toxins by using ribosome profiling, a new application of this technology. We found that toxin proteins evolve very quickly, and that they can change their tRNA targets during evolution. We also found that the corresponding immunity proteins evolve quickly.

Our results show that this class of tRNA-cleaving killer toxin pre-dates the genetic code changes in budding yeasts, so they are consistent with the hypothesis that the reassignment of the CUG codon was driven by a tRNA-cleaving killer toxin, although we did not find a toxin that specifically cleaves tRNA-Leu(CAG).

Results from the project were published as open-access papers in international peer-reviewed papers during 2022-2025.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

The results obtained have greatly expanded our knowledge of the tRNA-cleaving (anticodon nuclease) family of killer toxins, and showed that this family is highly heterogeneous and evolving very rapidly. Quite unexpectedly, we found that similar killer toxins are also made by filamentous fungi such as Fusarium, where the toxins and their immunity factors are encoded by small clusters of genes in the nuclear genome. We also found that cytosolic plasmids, similar to the cytosolic plasmids that encode killer toxins in budding yeasts, are present in basal fungi such as Coemansia species. Our results have given the field a much clearer understanding of the ubiquity of this class of killer toxin, and its role in the evolution of the genetic code.

fig-1-genetic-code.png