CRISPR/Cas9-Mediated Genome Engineering Reveals the Contribution of the 26S Proteasome to the Extremophilic Nature of the Yeast Debaryomyces hansenii.
ACS Synth Biol
; 10(2): 297-308, 2021 02 19.
Article
em En
| MEDLINE
| ID: mdl-33501828
ABSTRACT
The marine yeast Debaryomyces hansenii is of high importance in the food, chemical, and medical industries. D. hansenii is also a popular model for studying molecular mechanisms of halo- and osmotolerance. The absence of genome editing technologies hampers D. hansenii research and limits its biotechnological application. We developed novel and efficient single- and dual-guide CRISPR systems for markerless genome editing of D. hansenii. The single-guide system allows high-efficiency (up to 95%) mutation of genes or regulatory elements. The dual-guide system is applicable for efficient deletion of genomic loci. We used these tools to study transcriptional regulation of the 26S proteasome, an ATP-dependent protease complex whose proper function is vital for all cells and organisms. We developed a genetic approach to control the activity of the 26S proteasome by deregulation of its essential subunits. The mutant strains were sensitive to geno- and proteotoxic stresses as well as high salinity and osmolarity, suggesting a contribution of the proteasome to the extremophilic properties of D. hansenii. The developed CRISPR systems allow efficient D. hansenii genome engineering, providing a genetic way to control proteasome activity, and should advance applications of this yeast.
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Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Saccharomyces cerevisiae
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Complexo de Endopeptidases do Proteassoma
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Debaryomyces
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Sistemas CRISPR-Cas
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Edição de Genes
Tipo de estudo:
Prognostic_studies
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article