Hokuto Ohtsuka

A novel gene, ecl1+, extends the chronological lifespan in fission yeast

We have identified a novel gene from Schizosaccharomyces pombe that we have named ecl1(+) (extender of the chronological lifespan). When ecl1(+) is provided on a high-copy number plasmid, it extends the viability of both the Deltasty1 MAP kinase mutant and the wild-type cells after entry into the stationary phase. ecl1(+) encodes an 80-amino acid polypeptide that had not been annotated in the current database. The ecl1(+)-mRNA increases transiently when the growth phase is changed from the log phase to the stationary phase. The Ecl1 protein is localized in the nucleus. Calorie restriction extends the chronological lifespan of wild-type and Deltaecl1 cells but not ecl1(+)-overproducing cells. The Deltapka1 mutant shows little, if any, additional extension of viability when Ecl1 is overproduced. The ste11(+) gene that is negatively controlled by Pka1 is up regulated when Ecl1 is overproduced. From these results we propose that the effect of Ecl1 overproduction may be mainly linked to and negatively affects the Pka1-dependent pathway.

Identification of Ecl Family Genes That Extend Chronological Lifespan in Fission Yeast

In fission yeast, we identified two genes, named ecl2 + and ecl3 +, that are paralogous to ecl1 +, which extends the chronological lifespan. Both ecl2 + and ecl3 + extend the chronological lifespan when overexpressed as ecl1 +. ecl2 + and ecl3 + encode 84- and 89-amino acid polypeptides respectively that are not annotated in the current database. The Ecl2 protein is localized mainly in the nucleus, as Ecl1. These results suggest that ecl1 +, ecl2 +, and ecl3 + have overlapping functions in the regulation of chronological lifespan.

Pma1, a P-type Proton ATPase, Is a Determinant of Chronological Life Span in Fission Yeast

Chronological life span is defined by how long a cell can survive in a non-dividing state. In yeast, it is measured by viability after entry into stationary phase. To date, some factors affecting chronological life span have been identified; however, the molecular details of how these factors regulate chronological life span have not yet been elucidated clearly. Because life span is a complicated phenomenon and is supposedly regulated by many factors, it is necessary to identify new factors affecting chronological life span to understand life span regulation. To this end, we have screened for long-lived mutants and identified Pma1, an essential P-type proton ATPase, as one of the determinants of chronological life span. We show that partial loss of Pma1 activity not only by mutations but also by treatment with the Pma1 inhibitory chemical vanadate resulted in the long-lived phenotype in Schizosaccharomyces pombe. These findings suggest a novel way to manipulate chronological life span by modulating Pma1 as a molecular target.

hsf1+ extends chronological lifespan through Ecl1 family genes in fission yeast

The heat shock factor (HSF), a protein evolutionarily conserved from yeasts to human, regulates the expression of a set of proteins called heat shock proteins (HSPs), many of which function as molecular chaperones. In Saccharomyces cerevisiae, the HSF binds to the 5′ upstream region of YGR146C and activates its transcription. YGR146C encodes a functional homolog of ecl1+, ecl2+, and ecl3+ of Schizosaccharomyces pombe. At present, these Ecl1 family genes, which are extenders of chronological lifespan, have been identified only in fungi groups. Based on ChIP analysis, we identified that Hsf1 binds to the upstream DNA region of ecl2+ after heat shock in S. pombe. In Caenorhabditis elegans, heat shock factor HSF-1 is known to regulate aging and required for the elongation of longevity by dietary restriction. We found that heat shock factor Hsf1 extends chronological lifespan of S. pombe when overexpressed. Moreover, we show that the extension of chronological lifespan by the overproduction of Hsf1 mainly depends on ecl2+ among Ecl1 family genes. From these results, we suggest that HSF is a conserved regulator of lifespan, at least in yeast and nematode, and Ecl1 family genes such as YGR146C and ecl2+ are the direct targets of Hsf1 and mediate lifespan extension by Hsf1.

Chronological lifespan extension by Ecl1 family proteins depends on Prr1 response regulator in fission yeast.

ecl1 +, ecl2+ and ecl3+ genes encode highly homologous small proteins, and their over‐expressions confer both H2O2 stress resistance and chronological lifespan extension on Schizosaccharomyces pombe. However, the mechanisms of how these Ecl1 family proteins function have not been elucidated. In this study, we conducted microarray analysis and identified that the expression of genes involved in sexual development and stress responses was affected by the over‐expression of Ecl1 family proteins. In agreement with the mRNA expression profile, the cells over‐expressing Ecl1 family proteins showed high mating efficiency and resistant phenotype to H2O2. We showed that the H2O2‐resistant phenotype depends on catalase Ctt1, and over‐expression of ctt1+ does not affect chronological lifespan. Furthermore, we showed that six genes, ste11+, spk1+, hsr1+, rsv2+, hsp9+ and lsd90+, whose expressions are increased in cells over‐expressing Ecl1 family proteins are involved in chronological lifespan in fission yeast. Among these genes, the induction of ste11+ and hsr1+ was dependent on a transcription factor Prr1, and we showed that the extensions of chronological lifespan by Ecl1 family proteins are remarkably diminished in prr1 deletion mutant. From these results, we propose that Ecl1‐family proteins conduct H2O2 stress resistance and chronological lifespan extension in ctt1+‐ and prr1+‐dependent manner, respectively.

Identification of a Fatty Acyl-CoA Synthetase Gene, lcf2^+, Which Affects Viability after Entry into the Stationary Phase in Schizosaccharomyces pombe

The lcf1 + gene, which encodes a long chain fatty acyl-CoA synthetase, is necessary for the maintenance of viability after entry into the stationary phase in Schizosaccharomyces pombe. In this study, we analyzed a paralogous gene, SPBP4H10.11c (named lcf2 +), and we present evidence that the gene encodes a new fatty acyl-CoA synthetase. The enzyme preferentially recognized myristic acid as a substrate. A Δlcf2 mutant showed increased viability after entry into the stationary phase in SD medium. A Δlcf1Δlcf2 double mutant showed a severe decrease in long-chain fatty acyl-CoA synthetase activity and a rapid loss of viability after entry into the stationary phase. These results suggest that fatty acid utilization and/or metabolism is important to determine viability in the stationary phase.

Screening for long-lived genes identifies Oga1, a guanine-quadruplex associated protein that affects the chronological lifespan of the fission yeast Schizosaccharomyces pombe.

Schizosaccharomyces pombe and Saccharomyces cerevisiae are excellent model organisms to study lifespan. We conducted screening to identify novel genes that, when overexpressed, extended the chronological lifespan of fission yeast. We identified seven genes, among which we focused on SPBC16A3.08c. The gene product showed similarity to Ylr150w of S. cerevisiae, which has affinity for guanine-quadruplex nucleic acids (G4). The SPBC16A3.08c product associated with G4 in vitro and complemented the phenotype of an S. cerevisiae Ylr150w deletion mutant. From these results, we proposed that SPBC16A3.08c encoded for a functional homolog of Ylr150w, which we designated ortholog of G4-associated protein (oga1+). oga1+ overexpression extended the chronological lifespan and also decreased mating efficiency and caused both high and low temperature-sensitive growth. Deleting oga1+ resulted in caffeine-sensitive and canavanine-resistant phenotypes. Based on these results, we discuss the function of Oga1 on the chronological lifespan of fission yeast.

Sexual development of Schizosaccharomyces pombe is induced by zinc or iron limitation through Ecl1 family genes.

Ecl1 family genes (ecl1+, ecl2+, and ecl3+) have been identified as extenders of the chronological lifespan in Schizosaccharomyces pombe. Here, we found that the triple-deletion mutant (∆ecl1/2/3) had a defect in sexual development after entry into the stationary phase, although the mutant essentially showed normal mating and sporulation under nitrogen starvation or carbon limitation. In this study, we showed that limitation of zinc or iron can be a signal for sexual development of S. pombe cells grown in Edinburgh minimal medium until the stationary phase and that Ecl1 family genes are important for this process. Because the ∆ecl1/2/3 mutant diminishes the zinc depletion-dependent gene expression, Ecl1 family proteins may function as zinc sensors in the process of sexual development.

The Fission Yeast php2 Mutant Displays a Lengthened Chronological Lifespan

The Schizosaccharomyces pombe php2 + gene encodes a subunit of the CCAAT-binding factor complex. We found that disruption of the php2 + gene extended the chronological lifespan of the fission yeast. Moreover, the lifespan of the Δphp2 mutant was barely extended under calorie restricted (CR) conditions. Many other phenotypes of the Δphp2 mutant resembled those of wild-type cells grown under CR conditions, suggesting that the Δphp2 mutant might undergo CR. The mutant also showed low respiratory activity concomitant with decreased expression of the cyc1 + and rip1 + genes, both of which are involved in mitochondrial electron transport. On the basis of a chromatin immunoprecipitation assay, we determined that Php2 binds to a DNA region upstream of cyc1 + and rip1 + in S. pombe. Here we discuss the possible mechanisms by which the chronological lifespan of Δphp2 mutant is extended.

Ecl1, a regulator of the chronological lifespan of Schizosaccharomyces pombe, is induced upon nitrogen starvation.

In fission yeast, ecl1 + was identified as a novel factor that extends chronological lifespan when overexpressed. Ecl1 is a small protein consisting of 80 amino acids localized mainly in the nucleus. However, the mechanism by which it affects chronological lifespan has not been elucidated clearly. Here we analyzed the expression profile of Ecl1, especially as to cell cycle and growth phase, and found that it is induced upon nitrogen starvation. Then we analyzed the relevance of factors, Atf1, Ste11, and Tor1, which are known to be involved in the signaling of nitrogen starvation. Though the nitrogen starvation-induced expression of Ecl1 did not change in the atf1Δ mutant, induction in both the ste11Δ mutant and the tor1Δ mutant showed a delay. Based on these observations, the regulation of Ecl1 is discussed.