Codon usage bias (the preferential use of certain synonymous codons (optimal) over others is found at the organism level (intergenomic) within specific genomes (intragenomic) and even in certain genes. Whether it is the result of genetic drift due to GC/AT content and/or natural selection is a topic of intense debate. Preferential codons are mostly found in genes encoding highly-expressed proteins, while lowly-expressed proteins usually contain a high proportion of rare (lowly-represented) codons. While optimal codons are decoded by highly expressed tRNAs, rare codons are usually decoded by lowly-represented tRNAs. Whether rare codons play a role in controlling the expression of lowly- or temporarily-expressed proteins is an open question. In this work we approached this question using two strategies, either by replacing rare glycine codons with optimal counterparts in the gene that encodes the cell cycle protein Cdc13, or by overexpression the tRNAGly that decodes rare codons from the fission yeast, Schizosaccharomyces pombe. While the replacement of synonymous codons severely affected cell growth, increasing tRNA levels affected the aggregation status of Cdc13 and cell division. These lead us to think that rare codons in lowly-expressed cyclin proteins are crucial for cell division, and that the overexpression of tRNA that decodes rare codons affects the expression of proteins containing these rare codons. These codons may be the result of the natural selection of codons in genes that encode lowly-expressed proteins.
Brettanomyces bruxellensis is regarded as the main spoilage microorganism in the wine industry, owing to its production of off-flavours. It is difficult to eradicate owing to its high tolerance of adverse environmental conditions, such as low nutrient availability, low pH, and high levels of ethanol and SO2. In this study, the production of volatile phenols and the growth kinetics of isolates from various regions of Chile were evaluated under stressful conditions. Through randomly amplified polymorphic DNA (RAPD) analysis, 15 strains were identified. These were grown in the presence of p-coumaric acid, a natural antimicrobial and the main precursor of off-flavours, and molecular sulfur dioxide (mSO2), an antimicrobial synthetic used in the wine industry. When both compounds were used simultaneously, there were clear signs of an improvement in the fitness of most of the isolates, which showed an antagonistic interaction in which p-coumaric acid mitigates the effects of SO2. Fourteen strains were able to produce 4-vinylphenol, which showed signs of phenylacrylic acid decarboxylase activity, and most of them produced 4-ethylphenol as a result of active vinylphenol reductase. These results demonstrate for the first time the serious implications of using p-coumaric acid, not only for the production of off-flavours, but also for its protective action against the toxic effects of SO2.
Brettanomyces bruxellensis is considered the most significant contaminant yeast in the wine industry since it causes a deterioration in the organoleptic properties of the wine and significant economic losses. This deterioration is due to the production of volatile phenols from hydroxycinnamic acids. These compounds possess antimicrobial properties; however, B. bruxellensis can resist this effect because it metabolizes them into less toxic ones. Recent studies have reported that B. bruxellensis grows under different stress conditions, including p-coumaric acid (pCA) but effective methods for its control have not been found yet. Since that in other yeasts, such as Saccharomyces cerevisiae, it has been described that light affects its growth, and we evaluated whether the light would have a similar effect on B. bruxellensis. The results show that at light intensities of 2,500 and 4,000 lux in the absence of pCA, B. bruxellensis LAMAP2480 does not grow in the culture medium; however, when the medium contains this acid, the yeast adapts to both factors of stress managing to grow. The expression of genes related to oxidative stress in B. bruxellensis LAMAP2480, such as SOD1, GCN4, and ESBP6, showed a higher relative expression when the yeast was exposed to 2,500 lux compared to 4,000 lux, agreeing with the growth curves. This suggests that a higher expression of the genes studied would be related to stress-protective effects by pCA.
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