Die Bedeutung der universell konservierten ATPase YchF für die zelluläre Stressantwort
Zusammenfassung der Projektergebnisse
YchF/Ola1 represent universally conserved ATPases that have been implicated in multiple cellular processes, but their exact mode of action is still unknown. In a complementary approach, we analyzed the role of YchF/Ola1 in the prokaryotic model organism E. coli and the eukaryotic model organism S. cerevisiae. Our data demonstrate conserved traits but also variations between the prokaryotic YchF and the eukaryotic Ola1. Conserved traits include the ability to bind to ribosomes and its involvement in the cellular stress response. Unexpectedly, while the deletion of YchF in E. coli causes a gain-of-function phenotype under stress conditions, the absence of Ola1 in yeast is associated with a loss-of-function phenotype during stress. The gain-of-function phenotype in E. coli can be explained by the stationary-phase like condition of the ∆ychF strain. This is deduced from the observed increased levels of RpoS, which is the major transcriptional regulator that regulates the transition from exponential phase into stationary phase. The down-regulation of many metabolic processes and the simultaneous up-regulation of detoxifying enzymes, like KatE, make these cells less susceptible to inhibitors. Our data indicate that the RpoS increase and the increased catalase activity is the result of enhanced translation of the respective mRNAs, consolidating the data on ribosome binding and stress resistance. Thus, the current model proposes a role of YchF in selectively preventing the translation of those mRNAs that are specifically required only during stationary phase. Additionally, YchF appears to stimulate the formation of the signaling molecules ppGpp and polyP. Synthesis of ppGpp has been shown to occur by the ribosome-associated protein RelA, further supporting the model that YchF acts at the ribosome. In the yeast Saccharomyces cerevisiae, depletion of Ola1 resulted in a lossof-function phenotype under oxidative, heat and translational stress. Interestingly, our data show that Ola1 interacts with the ribosomes as well as co-localizes to stress granules upon severe heat stress. It is assumed that stress granules are a storage form of various proteins which are required for efficient restarting of cell cycle and growth after relief of stress. Accordingly, ∆ola1 cells exhibited a decreased growth during recovery from severe heat stress. Our data further show that loss of Ola1 does not alter stress granule formation but leads to increased levels of protein ubiquitination upon heat stress. Currently, we are proposing two hypotheses concerning the role of Ola1 in S. cerevisiae. On the one hand, Ola1 might exhibit chaperone or co-chaperone-like activity and is required to maintain intracellular protein homeostasis. Consistently, it has been reported that chaperones co-localize to heat stress granules and drive their disassembly. On the other hand, Ola1 might have an impact on inhibition of translation initiation upon stress which would be in line with previous findings in human cells where the GTPase activity of hOla1 interferes with ternary complex formation and results in suppression of global translation.
Projektbezogene Publikationen (Auswahl)
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(2016) Redox-activation of the universally conserved ATPase YchF by thioredoxin 1. Antioxidants and Redox Signaling, 24, 141-156
Hannemann, L., Suppanz, I., Ba, Q., MacInnes, K., Drepper, F., Warscheid, B., and Koch, H.G.
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(2017) Definition of a high-confidence mitochondrial proteome at quantitative scale. Cell Rep. 19, 2836-2852
Morgenstern, M., Stiller, S.B., Lübbert, P., Peikert, C.D., Dannenmaier, S., Drepper, F., Weill, U., Höß, P., Feuerstein, R., Gebert, M., Bohnert, M., van der Laan, M., Schuldiner, M., Schütze, C., Oeljeklaus, S., Pfanner, N., Wiedemann, N., Warscheid, B.
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(2018) Complete native stable isotope labeling by amino acids of Saccharomyces cerevisiae for global proteomic analysis. Anal Chem., 90, 10501-10509
Dannenmaier, S., Stiller, S.B., Morgenstern, M., Lübbert, P., Oeljeklaus, S., Wiedemann, N., Warscheid, B.
