Deubiquitinating activity of Sdu1, a putative member of the PPPDE peptidase family, inSchizosaccharomyces pombe

Abstract

The Schizosaccharomyces pombe sdu⁺ gene encoding a putative member of the PPPDE (Permuted Papain fold Peptidases of DsRNA viruses and Eukaryotes) superfamily was cloned into an Escherichia coli - yeast shuttle vector pRS316, resulting in the recombinant plasmid pYSTP. The determined nucleotide sequence carries 1207 bp, which would encode a protein of 201 amino acid residues. The S. pombe cells harboring pYSTP contained higher sdu1⁺ mRNA and deubiquitinating activity levels than the vector control cells, indicating that the sdu1⁺ gene is functioning. They exhibited a better growth in normal rich medium than the vector control cells. When shifted into the fresh medium containing hydrogen peroxide, menadione, or sodium nitroprusside, the S. pombe cells harboring pYSTP were able to grow reasonably well, while the growth of the vector control cells was arrested. The reactive oxygen species and total glutathione levels of the S. pombe cells harboring pYSTP were lower and higher than those of the vector control cells under the same stressful conditions, respectively. They exhibited a lower nitric oxide level than the vector control cells when subjected to sodium nitroprusside. Taken together, the sdu1⁺ gene encodes an actual protein having deubiquitinating activity and is involved in the response against oxidative and nitrosative stresses in S. pombe.

Reactive oxygen species-dependent down-regulation of ubiquitin C-terminal hydrolase in Schizosaccharomyces pombe

“…Next, changes in the UCH activities, encoded by the sdu1 + gene, were compared in the Sdu1-overexpressing S. pombe cells and the vector control cells under stress conditions. As shown in the previous report (Kim et al, 2013), the S. pombe cells harboring pYSTP were found to contain the UCH activity 3.4-fold higher than the vector control cells (Fig. 4; open and closed bars, lane 1).…”

“…+ gene, known as a member of the PPPDE peptidase superfamily of DUBs (Iyer et al, 2004), was cloned from the fission yeast Schizosaccharomyces pombe and shown to encode an UCH activity (Kim et al, 2013). Sdu1 is regarded as the first example of a PPPDE superfamily member with UCH activity.…”

“…Sdu1 is regarded as the first example of a PPPDE superfamily member with UCH activity. It was confirmed to participate in the response against oxidative and nitrosative stresses in S. pombe by modulating the reactive oxygen species (ROS) and total glutathione (GSH) levels (Kim et al, 2013 , was used in this work. The Sdu1-overexpressing recombinant plasmid pYSTP was previously constructed using a yeast-E. coli shuttle plasmid vector pRS316 (Myers et al, 1986).…”

Defensive roles of Sdu1, a PPPDE superfamily member with ubiquitin C-terminal hydrolase activity, against thermal stress in Schizosaccharomyces pombe

“…Key words: Schizosaccharomyces pombe, glutathione, reactive oxygen species, Sdu1, thermal stress In the previous work, the sdu1 + gene encoding Sdu1, belonging to the PPPDE superfamily, was cloned from S. pombe, and its gene product was shown to contain the UCH activity which is involved in the defense against oxidative and nitrosative stresses in S. pombe (Kim et al, 2013). In the present work, we further demonstrate that Sdu1 plays a defensive role against thermal stress in S. pombe.…”

“…Total superoxide dismutase (SOD) and GSH peroxidase activities were also higher in the S. pombe cells harboring pYSTP after the temperature shifts than in the vector control cells. In brief, the S. pombe Sdu1 plays a thermotolerant role against high incubation temperature through the down-regulation of ROS and NO and the up-regulation of total GSH content, total SOD and GSH peroxidase activities.Key words: Schizosaccharomyces pombe, glutathione, reactive oxygen species, Sdu1, thermal stress In the previous work, the sdu1 + gene encoding Sdu1, belonging to the PPPDE superfamily, was cloned from S. pombe, and its gene product was shown to contain the UCH activity which is involved in the defense against oxidative and nitrosative stresses in S. pombe (Kim et al, 2013). In the present work, we further demonstrate that Sdu1 plays a defensive role against thermal stress in S. pombe.…”