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A lncRNA-regulated gene expression system with rapid induction kinetics in the fission yeast Schizosaccharomyces pombe.

ABSTRACT: The fission yeast Schizosaccharomyces pombe is an excellent model organism for the study of eukaryotic cellular physiology. The organism is genetically tractable and several tools to study the functions of individual genes are available. One such tool is regulatable gene expression and overproduction of proteins. Limitations of currently available overexpression systems include delay in expression after induction, narrow dynamic range, and system-wide changes due to induction conditions. Here I describe a new long noncoding RNA (lncRNA)-regulated, thiamine-inducible expression system that integrates lncRNA-based transcriptional interference at the fission yeast tgp1 promoter with the fast repression kinetics of the thiamine-repressible nmt1 promoter. This hybrid system has rapid induction kinetics, broad dynamic range, and tunable expression via thiamine concentration. The lncRNA-regulated thiamine-inducible system will be advantageous for the study of individual genes and for potential applications in the production of heterologous proteins in fission yeast.

SUBMITTER: Garg A

PROVIDER: S-EPMC7566572 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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A lncRNA-regulated gene expression system with rapid induction kinetics in the fission yeast <i>Schizosaccharomyces pombe</i>.

Garg Angad A

RNA (New York, N.Y.) 20200811 11

The fission yeast <i>Schizosaccharomyces pombe</i> is an excellent model organism for the study of eukaryotic cellular physiology. The organism is genetically tractable and several tools to study the functions of individual genes are available. One such tool is regulatable gene expression and overproduction of proteins. Limitations of currently available overexpression systems include delay in expression after induction, narrow dynamic range, and system-wide changes due to induction conditions. ...[more]

PMID: 32788323

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Project description:BackgroundHIV-1 protease (PR) is an essential viral enzyme. Its primary function is to proteolyze the viral Gag-Pol polyprotein for production of viral enzymes and structural proteins and for maturation of infectious viral particles. Increasing evidence suggests that PR cleaves host cellular proteins. However, the nature of PR-host cellular protein interactions is elusive. This study aimed to develop a fission yeast (Schizosaccharomyces pombe) model system and to examine the possible interaction of HIV-1 PR with cellular proteins and its potential impact on cell proliferation and viability.ResultsA fission yeast strain RE294 was created that carried a single integrated copy of the PR gene in its chromosome. The PR gene was expressed using an inducible nmt1 promoter so that PR-specific effects could be measured. HIV-1 PR from this system cleaved the same indigenous viral p6/MA protein substrate as it does in natural HIV-1 infections. HIV-1 PR expression in fission yeast cells prevented cell proliferation and induced cellular oxidative stress and changes in mitochondrial morphology that led to cell death. Both these PR activities can be prevented by a PR-specific enzymatic inhibitor, indinavir, suggesting that PR-mediated proteolytic activities and cytotoxic effects resulted from enzymatic activities of HIV-1 PR. Through genome-wide screening, a serine/threonine kinase, Hhp2, was identified that suppresses HIV-1 PR-induced protease cleavage and cell death in fission yeast and in mammalian cells, where it prevented PR-induced apoptosis and cleavage of caspase-3 and caspase-8.ConclusionsThis is the first report to show that HIV-1 protease is functional as an enzyme in fission yeast, and that it behaves in a similar manner as it does in HIV-1 infection. HIV-1 PR-induced cell death in fission yeast could potentially be used as an endpoint for mechanistic studies, and this system could be used for developing a high-throughput system for drug screenings.

Dataset Information

A lncRNA-regulated gene expression system with rapid induction kinetics in the fission yeast Schizosaccharomyces pombe.

Publications

A lncRNA-regulated gene expression system with rapid induction kinetics in the fission yeast <i>Schizosaccharomyces pombe</i>.

Similar Datasets

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