| Popis: |
Reactive oxygen species (ROS) are byproduct of bacterial aerobic metabolism, however, if the metabolism is disrupted, amount of ROS increases beyond cell's capacity which causes oxidative stress. Additionally, oxidative stress can be caused by other bacteria that produce and excrete H2O2 or by phagocytes that produce ROS as a defence mechanism against pathogenic bacteria. Interestingly, it was shown in some cases that one stress can preadapt cells to other stress by activating stress response. Here, we investigated the effect of specific mistranslation of isoleucine with structurally similar amino acids valine (Val) and norvaline (Nva), on bacterial response to oxidative stress. We used Escherichia coli strain expressing mutant variant of isoleucyl-tRNA synthetase with inactivated editing domain which is more prone to mistranslation compared to wild type. Variable mistranslation rates were induced by different concentrations of Val or Nva added in the medium and oxidative stress was induced by adding H2O2. We observed reduced growth rate of bacteria in the media supplemented with Val or Nva and prolonged lag phase in the media supplemented with H2O2. However, prior exposure to mistranslation stress shortened the lag phase in oxidative stress after which bacteria continued to grow normally. Survival assays resulted in low survival rate of nonmistranslated culture in oxidative stress, while survival rates of cultures preexposed to mistranslation stress were higher. Using light microscopy, we observed filamentation of bacteria. Under transmission electron microscope, bacterial cells grown with 1 mM Val or Nva exhibited profound morphological and ultrastructural changes, such as numerous intracellular vesicles. More pronounced filamentation of bacteria grown with higher Val or Nva concentrations implies stronger stress response activation which correlates with better proliferation and viability. The results suggest that mistranslation, specifically, mistranslation of Ile by Val and Nva, induces cellular response that increases bacterial tolerance to oxidative stress. The further research in proteomics is planned to elucidate cellular mechanisms of adaptative response induced by mistranslation. |
