The Schizosaccharomyces pombe transcription factor Pap1 regulates antioxidant-gene transcription in response to H ₂ O ₂ . Pap1 activation occurs only at low, but not elevated, H ₂ O ₂ concentrations that instead strongly trigger the mitogen-activated protein kinase Sty1 pathway. Here, we identify the peroxiredoxin Tpx1 as the upstream activator of Pap1. We show that, at low H ₂ O ₂ concentrations, this oxidant scavenger can transfer a redox signal to Pap1, whereas higher concentrations of the oxidant inhibit the Tpx1-Pap1 redox relay through the temporal inactivation of Tpx1 by oxidation of its catalytic cysteine to a sulfinic acid. This cysteine modification can be reversed by the sulfiredoxin Srx1, its expression in response to high doses of H ₂ O ₂ strictly depending on active Sty1. Thus, Tpx1 oxidation to the cysteine-sulfinic acid and its reversion by Srx1 constitutes a previously uncharacterized redox switch in H ₂ O ₂ signaling, restricting Pap1 activation within a narrow range of H ₂ O ₂ concentrations.

The Elongator complex, including the histone acetyl transferase Sin3/Elp3, was isolated as an RNA polymerase II-interacting complex, and cells deficient in Elongator subunits display transcriptional defects. However, it has also been shown that Elongator mediates the modification of some tRNAs, modulating translation efficiency. We show here that the fission yeast Sin3/Elp3 is important for oxidative stress survival. The stress transcriptional program, governed by the Sty1-Atf1-Pcr1 pathway, is affected in mutant cells, but not severely. On the contrary, cells lacking Sin3/Elp3 cannot modify the uridine wobble nucleoside of certain tRNAs, and other tRNA modifying activities such as Ctu1-Ctu2 are also essential for normal tolerance to H₂O₂. In particular, a plasmid over-expressing the tRNA^Lys_UUU complements the stress-related phenotypes of Sin3/Elp3 mutant cells. We have determined that the main H₂O₂-dependent genes, including those coding for the transcription factors Atf1 and Pcr1, are highly expressed mRNAs containing a biased number of lysine-coding codons AAA versus AAG. Thus, their mRNAs are poorly translated after stress in cells lacking Sin3/Elp3 or Ctu2, whereas a mutated atf1 transcript with AAA-to-AAG lysine codons is efficiently translated in all strain backgrounds. Our study demonstrates that the lack of a functional Elongator complex results in stress phenotypes due to its contribution to tRNA modification and subsequent translation inefficiency of certain stress-induced, highly expressed mRNAs. These results suggest that the transcriptional defects of these strain backgrounds may be a secondary consequence of the deficient expression of a transcription factor, Atf1-Pcr1, and other components of the transcriptional machinery. The success of a biological event such as cellular adaptation to environmental changes requires the complex process of protein expression to be carried out with high efficiency and fidelity. Thus, not only transcription but also mRNA homeostasis and translation have to be performed with maximum efficiency, or survival would be hampered. Our study demonstrates that the role of Elongator, a putative Pol II-associated complex, in survival to stress is to optimize translation efficiency by modifying some particular tRNAs. We show here that Sin3/Elp3, an Elongator component, participates in the modification of the anticodon of the low copy number tRNA^Lys_UUU, which probably favours codon recognition. This tRNA recognizes one of the two codons for lysine, which is down-represented in highly expressed constitutive genes. The stress mRNAs, highly-expressed upon stress conditions, have not adapted their lysine codon usage from AAA-to-AAG, and proper tRNA^Lys_UUU modification by Elongator is an alternative strategy to accomplish efficient translation of these AAA-containing, abundant stress mRNAs.

Either calorie restriction, loss‐of‐function of the nutrient‐dependent PKA or TOR/SCH9 pathways, or activation of stress defences improves longevity in different eukaryotes. However, the molecular links between glucose depletion, nutrient‐dependent pathways and stress responses are unknown. Here, we show that either calorie restriction or inactivation of nutrient‐dependent pathways induces lifespan extension in fission yeast, and that such effect is dependent on the activation of the stress‐dependent Sty1 mitogen‐activated protein (MAP) kinase. During transition to stationary phase in glucose‐limiting conditions, Sty1 becomes activated and triggers a transcriptional stress programme, whereas such activation does not occur under glucose‐rich conditions. Deletion of the genes coding for the SCH9‐homologue, Sck2 or the Pka1 kinases, or mutations leading to constitutive activation of the Sty1 stress pathway increase lifespan under glucose‐rich conditions, and importantly such beneficial effects depend ultimately on Sty1. Furthermore, cells lacking Pka1 display enhanced oxygen consumption and Sty1 activation under glucose‐rich conditions. We conclude that calorie restriction favours oxidative metabolism, reactive oxygen species production and Sty1 MAP kinase activation, and this stress pathway favours lifespan extension.

Summary: The transcription factor Pap1 and the MAP kinase Sty1 are key regulators of hydrogen peroxide‐induced responses in Schizosaccharomyces pombe. Pap1 can be activated quickly at low, but not high, hydrogen peroxide concentrations. The MAP kinase Sty1 has been reported to participate in Pap1 activation by the oxidant. Here, we provide biochemical and genetic evidence for the in vivo formation of a hydrogen peroxide‐induced disulphide bond in Pap1, which precedes the rapid and reversible nuclear accumulation of the transcription factor. We show that activation of the Sty1 cascade before the oxidative insult, or overexpression of the Sty1‐regulated genes ctt1 (encoding catalase) or gpx1 (encoding glutathione peroxidase), can accelerate Pap1 entry even at high doses of hydrogen peroxide. In fact, the lack of Sty1 impedes Pap1 nuclear localization, but only at high doses of the oxidant. We propose that, whereas low doses of hydrogen peroxide lead directly to Pap1 oxidation‐activation, high concentrations of the oxidant initially activate the Sty1 pathway, with the consequent increase in scavenging enzymes, which in turn helps to decompose the excess of hydrogen peroxide and achieve an appropriate concentration for the subsequent activation of Pap1. Our results also suggest that activation of Sty1 at high doses of hydrogen peroxide may also be required to trigger other antioxidant activities such as those reverting the overoxidation of cysteine residues at the Pap1 pathway.