Thymol antifungal mode of action involves telomerase inhibition

The antifungal mode of action of thymol was investigated by a chemical-genetic profile analysis. Growth of each of ~4700 haploid Saccharomyces cerevisiae gene deletion mutants was monitored on medium with a subinhibitory concentration (50 μg/ml) of thymol and compared to growth on non-thymol control medium. This analysis revealed that, of the 76 deletion mutants with the greatest degree of susceptibility to thymol, 29% had deletions in genes involved in telomere length maintenance. A telomere restriction fragment (TRF) length assay showed that yeast exposed to a subinhibitory concentration of thymol for 15 days had telomere size reductions of 13-20% compared to non-thymol controls. By accelerating telomere shortening, thymol may increase the rate of cell senescence and apoptosis. Furthermore, real-time RT-PCR analysis revealed approximately two-fold reductions in EST2 mRNA but no change in TLC1 RNA in thymol-treated S. cerevisiae relative to untreated cells. EST2 encodes the essential reverse transcriptase subunit of telomerase that uses TLC1 RNA as a template during addition of TG(1-3) repeats to maintain telomere ends. This study provides compelling evidence that a primary mode of thymol antifungal activity is through inhibition of transcription of EST2 and thus telomerase activity.