A second protein disulfide isomerase plays a protective role against nitrosative and nutritional stresses in Schizosaccharomyces pombe

Abstract

In the present work, a second gene encoding protein disulfide isomerase (PDI2) was cloned and characterized from Schizosaccharomyces pombe, and its regulation was studied. The structural gene encoding PDI2 was amplified from the genomic DNA using PCR, and ligated into the E. coli-yeast shuttle vector pRS316 to generate the recombinant plasmid pYPDI2. The determined DNA sequence carries 2,578 bp and is able to encode a protein of 726 amino acid sequence with CGAC at the putative active site. The fission yeast cells harboring pYPDI2 contained 1.62- and 2.73-fold higher PDI activity than the control yeast cells in exponential and stationary phases, respectively, indicating that the cloned gene is in vivo functioning. The PDI2 mRNA levels in both vector control and pYPDI2-containing yeast cells were found to be significantly higher in the stationary phase than in the exponential phase, suggesting that expression of the PDI2 gene is under stationary control. The yeast cells harboring pYPDI2 showed enhanced survival on minimal media plates containing nitric oxide (NO)-generating sodium nitroprusside (SNP) and no nitrogen. The synthesis of β-galactosidase from the PDI2-lacZ fusion gene was markedly enhanced in the Pap1-positive KP1 cells by SNP and nitrogen starvation. However, the enhancement in the synthesis of β-galactosidase from the PDI2-lacZ fusion gene by SNP and nitrogen starvation appeared to be relatively reduced in the Pap1-negative TP108-3C cells than in the Pap1-positive KP1 cells. The PDI2 mRNA level was elevated by SNP and nitrogen starvation in the Pap1-positive cells but not in the Pap1-negative cells. In brief, the S. pombe PDI2 plays a protective role against nitrosative and nutritional stresses, and is positively regulated by NO and nitrogen starvation in a Pap1-dependent manner.

A unique thioredoxin reductase plays defensive roles against oxidative, nitrosative and nutritional stresses in Schizosaccharomyces pombe

“…Similarly, overexpression of a second protein disulfide isomerase, PDI2, increases S. pombe survival on nitrogendepleted minimal medium plates supplemented with NOgenerating SNP. In addition, S. pombe Pdi2 is positively regulated by NO and nitrogen starvation in a Pap1-dependent fashion (Lee et al, 2010). Our work also demonstrates that TrxR is involved in the response to nitrogen starvation in S. pombe.…”

Participation of protein disulfide isomerase 2 in the tolerance against mercury toxicity in Schizosaccharomyces pombe

“…The Aspergillus niger gene encoding PDI is also induced by ER stress (Jeenes et al, 1997). Both the S. pombe Pdi1 and Pdi2 were previously verified to participate in the control of cellular growth and response to nutritional and oxidative stresses (Kim et al, 2006;Lee et al, 2010). This work additionally demonstrates that the S.…”

“…pombe KP1 (h + leu1-32 ura4-294), used in the present work, was a derivative of S. pombe heterothallic haploid strain 975h + . The Pdi2-overexpressing recombinant plasmid pYPDI2 was previously constructed using a yeast-E. coli shuttle plasmid vector pRS316 (Lee et al, 2010). Yeast cells were grown in YEPD medium (pH 6.5) which contains 1% yeast extract, 2%…”

“…Through the previous works (Kim et al, 2006;Lee et al, 2010), the two isoforms of PDIs were cloned and basically characterized from the fission yeast Schizosaccharomyces pombe. The first PDI, later named Pdi1, which is very homologous to Arabidopsis thaliana PDI, is involved in the yeast growth, and its expression is up-regulated by oxidative stress, such as superoxide anion and hydrogen peroxide, and nitrogen starvation in a Pap1-independent manner (Kim et al, 2006).…”

“…The first PDI, later named Pdi1, which is very homologous to Arabidopsis thaliana PDI, is involved in the yeast growth, and its expression is up-regulated by oxidative stress, such as superoxide anion and hydrogen peroxide, and nitrogen starvation in a Pap1-independent manner (Kim et al, 2006). Pdi2, a second S. pombe PDI, plays a protective role against nitrosative and nutritional stresses, and its expression is positively regulated by nitric oxide (NO) and nitrogen starvation in a Pap1-dependent manner (Lee et al, 2010). In the present work using the Pdi2-overexpressing yeast cells, we demonstrate that Pdi2 participates in the tolerant response against mercury toxicity in S. pombe, possibly via specifically up-regulating total GSH and superoxide dismutase (SOD), but not glutathione peroxidase (GPx).…”