Regulation of the fission yeast transcription factor Pap1 by oxidative stress: requirement for the nuclear export factor Crm1 (Exportin) and the stress-activated MAP kinase Sty1/Spc1

Abstract

The fission yeast Sty1 stress-activated MAP kinase is crucial for the cellular response to a variety of stress conditions. Accordingly, sty1 − cells are defective in their response to nutrient limitation, lose viability in stationary phase, and are hypersensitive to osmotic stress, oxidative stress, and UV treatment. Some of these phenotypes are caused by Sty1-dependent regulation of the Atf1 transcription factor, which controls both meiosis-specific and osmotic stress-responsive genes. However, in this report we demonstrate that the cellular response to oxidative stress and to treatment with a variety of cytotoxic agents is the result of Sty1 regulation of the Pap1 transcription factor, a bZip protein with structural and DNA binding similarities to the mammalian c-Jun protein. We show that both Sty1 and Pap1 are required for the expression of a number of genes involved in the oxidative stress response and for the expression of two genes, hba2 + /bfr1 + and pmd1 + , which encode energy-dependent transport proteins involved in multidrug resistance. Furthermore, we demonstrate that Pap1 is regulated by stress-dependent changes in subcellular localization. On imposition of oxidative stress, the Pap1 protein relocalizes from the cytoplasm to the nucleus in a process that is dependent on the Sty1 kinase. This relocalization is the result of regulated protein export, rather than import, and involves the Crm1 (exportin) nuclear export factor and the dcd1 + /pim1 + gene that encodes an Ran nucleotide exchange factor.

The Peroxiredoxin Repair Proteins

“…Pap1 is an AP-1-like transcription factor that under normal conditions associates with the nuclear export factor Crm1 in the cytoplasm (Toone et al, 1998). Sty1 is a mitogen-activated protein kinase (MAPK) that is known to phosphorylate the transcription factor Atf1 (Shiozaki et al, 1995;Degols et al, 1996;Shiozaki et al, 1996;Wilkinson et al, 1996).…”

“…Sty1 is a mitogen-activated protein kinase (MAPK) that is known to phosphorylate the transcription factor Atf1 (Shiozaki et al, 1995;Degols et al, 1996;Shiozaki et al, 1996;Wilkinson et al, 1996). Under stress conditions with low levels of H 2 O 2 , Pap1 is activated by the formation of an intramolecular disulfide bond that prevents association with Crm1 and results in the nuclear accumulation (Toone et al, 1998;Kudo et al, 1999;Castillo et al, 2002;Quinn et al, 2002). In contrast, elevated levels of peroxide prevent nuclear accumulation and the expression of Pap1-dependent genes (Quinn et al, 2002;Vivancos et al, 2004).…”

Stress‐activated signalling pathways in yeast

“…Disruption of sty1 þ and wis1 þ results in a plethora of phenotypes all of which are related to the response of the cells to different stress conditions. Thus, sty1 ¹ cells are sensitive to osmotic, oxidative and heat stress, DNA damaging agents such as UV irradiation and cisplatin and to treatment with multiple drugs and heavy metals (Millar et al 1995;Shiozaki & Russell 1995;Kato et al 1996;Degols & Russell 1997;Toone et al 1998); in addition, they are deficient in initiating and progressing down the sexual differentiation pathway which results in spore formation and rapidly lose viability when the stationary phase is reached (Millar et al 1995;Shiozaki & Russell 1995;Kato et al 1996). These latter two phenotypes also reflect defects in stress response-in this case stress that arises from nutrient depletion.…”

“…A closer homologue to Pap1 is the budding yeast yAP1 factor that not only shares considerable sequence similarity within the DNA binding region but also within a cysteine rich domain (CRD) in the Cterminus, which is crucial for the regulation of Pap1 and yAP1 activity (see below). Deletion of Pap1 is not lethal, but it does result in cells that are very sensitive to a range of drugs and heavy metal treatment and to oxidative stress conditions (Toda et al 1991;Kumada et al 1996;Toone et al 1998). These are phenotypes shared by sty1 ¹ cells but not by cells devoid of atf1 þ .…”

“…Such transporters are known to be involved in multidrug resistance in many different eukaryotic organisms; in humans, they are very important at the clinical level since increased expression of such genes gives rise to multidrug resistant cells (Gottesman & Pastan 1993). The expression of both hba2 þ /bfr1 þ and pmd1 þ requires Pap1 and the Sty1 pathway (Toone et al 1998). With regard to the oxidative stress response, many genes have been described which encode proteins that defend against the potential harmful effects of ROS, and, in a number of eukaryotic organisms, these genes are upregulated.…”