Applied Stress Microbiology

Outline of Research and Education

Our research involves “Applied Molecular Microbiology”. Our laboratory aims at basic studies in microbial science, particularly cellular response and adaptation to environmental stresses, and practical applications in new biotechnology. To fully understand microbial cell functions under stress conditions, we analyze and improve various mechanisms of microorganisms from molecular, metabolic and cellular aspects. Novel findings from our basic studies may be applied to the molecular breeding of useful microbes (yeasts, bacteria), the production of valuable biomaterials (enzymes, amino acids) and the development of promising technologies to solve environmental issues (bioethanol, etc.).

To achieve these goals, we employ research methods in screening, genetics, biochemistry, molecular biology, enzymology, cell biology, post-genomic analysis, protein engineering, metabolic engineering and cell engineering based on the recent knowledge.

Major Research Topics

Stress response and tolerance in yeast Saccharomyces cerevisiae (Figs.1, 2, 3, 4)

We are interested in cellular response and adaptation to environmental stresses in the yeast Saccharomyces cerevisiae, which is an important microorganism as a model for higher eukaryotes. Yeast is also a useful microbe in the fermentation industry for the production of breads, alcoholic beverages and bioethanol. During fermentation, yeast cells are exposed to various stresses, including ethanol, high temperature, desiccation and osmotic pressure. Such stresses induce protein denaturation, reactive oxygen species generation, and lead to growth inhibition or cell death. In terms of application, stress tolerance is the key for yeast cells. We analyze the novel stress-tolerant mechanisms found in yeast listed below.

Proline: physiological functions, metabolic regulation, transport mechanisms

In response to osmotic stress, proline is accumulated in many bacterial and plant cells as an osmoprotectant. Previously, we found that proline is a very good cryoprotectant to yeast cells. We currently study the physiological functions and the transport mechanism of proline, and the functions of key enzymes in proline metabolism, γ-glutamyl kinase and proline oxidase.

N-Acetyltransferase Mpr1: arginine biosynthesis, antioxidative mechanisms

The Mpr1 gene found in S. cerevisiae Σ1278 encodes a novel N-acetyltransferase that detoxifies the proline analogue azetidine-2-carboxylate. We found that Mpr1 is involved in a novel pathway of arginine biosynthesis and protects yeast cells by regulating ROS levels under oxidative stress conditions. We will clarify the mechanism and physiological role of Mpr1-mediated arginine biosynthesis in yeast.

Nitric oxide (NO): synthetic mechanism, physiological roles

Nitric oxide (NO) produced by NO synthase (NOS) from arginine is a signaling molecule participates in many biological processes in mammals, but is poorly understood in yeast due to the lack of mammalian NOS orthologues. We recently discovered that NO is generated through the flavoprotein Tah18-dependent NOS-like activity and confers oxidative stress tolerance on yeast cells. We now analyze the synthetic mechanism and physiological roles of NO in yeast.

Ubiquitin (Ub) system: plasma membrane protein quality control, Ub ligase Rsp5 regulation.

We are also studying the ubiquitin (Ub) system involving Rsp5, an essential E3 ubiquitin ligase that governs many biological events. We showed that Rsp5 plays pivotal roles in both repair and degradation of stress-induced abnormal proteins. In particular, we hypothesize that Rsp5 is involved in selective ubiquitination for quality control of plasma membrane proteins under stress conditions.

Development of industrial yeast based on novel stress-tolerant mechanisms (Fig. 5)

Through our basic research on novel stress-tolerant mechanisms, we construct industrial yeasts with higher fermentation ability under various stress conditions and contribute to yeast-based industries for the effective production of bread dough and alcoholic beverages or breakthrough in bioethanol production.

Metabolic regulation and physiological roles of cysteine in Escherichia coli (Fig. 6)

Cysteine is an important amino acid both biologically and commercially. For achieving cysteine fermentation directly from glucose, we analyze the physiological roles and regulatory mechanisms of its biosynthesis, degradation and transport in Escherichia coli, based on genomic information and metabolic engineering.


Stress response and tolerance in yeast Saccharomyces cerevisiae

  1. Tatehashi Y. et al., FEBS Lett., in press
  2. Yoshikawa Y. et al., Nitric Oxide-Biol. Chem., 57, 85-91, 2016
  3. Astuti R. I. et al., Nitric Oxide-Biol. Chem., 52, 29-40, 2016
  4. Nasuno R. et al., J. Biochem., 159, 271-277, 2016
  5. Watanabe D. et al., Biochem. Biophys. Res. Commun., 463, 76-81, 2015
  6. Wijayanti I. et al., J. Biochem., 157, 251-260, 2015
  7. Nasuno R. et al., PLoS One, 9, e113788, 2014
  8. Oshiro S. & Takagi H., FEMS Yeast Res., 14, 1015-1027, 2014
  9. Shiga T. et al., Eukaryot. Cell, 13, 1191-1199, 2014
  10. Nasuno R. et al., Proc. Natl. Acad. Sci. USA, 110, 11821-11826, 2013
  11. Nishimura A. et al., Biochem. Biophys. Res. Commun., 430, 137-143, 2013
  12. Sasaki T. & Takagi H. Gene Cells, 18, 459-475, 2013
  13. Nishimura A. et al., FEMS Yeast Res., 10, 687-698, 2010
  14. Nomura M. and Takagi H., Proc. Natl. Acad. Sci. USA, 101, 12616-12621, 2004
  15. Hoshikawa C. et al., Proc. Natl. Acad. Sci. USA, 100, 11505-11510, 2003

Development of industrial yeast based on novel stress-tolerant mechanisms

  1. Watanabe D. et al., Appl. Environ. Microbiol., 82, 340-351, 2016
  2. Takagi H. et al., J. Biosci. Bioeng., 119, 140-147, 2015
  3. Watanabe D. et al., Appl. Environ. Microbiol., 78, 4008-4016, 2012
  4. Sasano Y. et al., Int. J. Food Microbiol., 152, 40-43 , 2012
  5. Sasano Y. et al., Microb. Cell Fact., 11:40 doi:10.1186/1475-2859-11-40, 2012

Physiological roles and metabolic regulation of cysteine in Escherichia coli

  1. Funahashi E. et al., J. Biosci. Bioeng., 120, 666-669, 2015
  2. Ohtsu I. et al., PLoS One, 10, e0120619, 2015
  3. Nakatani T. et al., Microb. Cell Fact., 11:62 doi:10.1186/1475-2859-11-62, 2012
  4. Ohtsu I. et al., J. Biol. Chem., 285, 17479-17487, 2010
  5. Wiriyathanawudhiwong et al., Appl. Microbiol. Biotech., 81, 903-913, 2009
Fig.1 Novel stress-tolerant mechanisms in S. cerevisiae.
Fig.2 Metabolic pathway of proline and arginine in S. cerevisiae.
Fig.3 NO-mediated antioxidative mechanism in S. cerevisiae.
Fig.4 Ubiquitin system under stress conditions in S. cerevisiae.
Fig.5 During fermentation processes, yeast cells are exposed to various stresses.
Fig.6 Sulfur metabolism in E. coli.
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