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New regulators of t...
New regulators of the tetracycline‐inducible gene expression system identified by chemical and genetic screens
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- Colicchia, Valeria (författare)
- Science for Life Laboratory, Division of Genome Biology, Department of Medical Biochemistry and Biophysics Karolinska Institute Stockholm Sweden
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- Häggblad, Maria (författare)
- Karolinska Institutet,Science for Life Laboratory, Division of Genome Biology, Department of Medical Biochemistry and Biophysics Karolinska Institute Stockholm Sweden
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- Sirozh, Oleksandra (författare)
- Genomic Instability Group Spanish National Cancer Research Centre (CNIO) Madrid Spain
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- Porebski, Bartlomiej (författare)
- Karolinska Institutet,Science for Life Laboratory, Division of Genome Biology, Department of Medical Biochemistry and Biophysics Karolinska Institute Stockholm Sweden
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- Balan, Mirela (författare)
- Science for Life Laboratory, Division of Genome Biology, Department of Medical Biochemistry and Biophysics Karolinska Institute Stockholm Sweden
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- Li, Xuexin (författare)
- Science for Life Laboratory, Division of Genome Biology, Department of Medical Biochemistry and Biophysics Karolinska Institute Stockholm Sweden
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- Lidemalm, Louise (författare)
- Science for Life Laboratory, Division of Genome Biology, Department of Medical Biochemistry and Biophysics Karolinska Institute Stockholm Sweden
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- Carreras-Puigvert, Jordi (författare)
- Uppsala universitet,Institutionen för farmaceutisk biovetenskap,Science for Life Laboratory, SciLifeLab
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- Hühn, Daniela (författare)
- Karolinska Institutet,Science for Life Laboratory, Division of Genome Biology, Department of Medical Biochemistry and Biophysics Karolinska Institute Stockholm Sweden
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- Fernandez‐Capetillo, Oscar (författare)
- Science for Life Laboratory, Division of Genome Biology, Department of Medical Biochemistry and Biophysics Karolinska Institute Stockholm Sweden;Genomic Instability Group Spanish National Cancer Research Centre (CNIO) Madrid Spain
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(creator_code:org_t)
- 2022-09-11
- 2022
- Engelska.
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Ingår i: FEBS Open Bio. - : Wiley-Blackwell. - 2211-5463. ; 12:10, s. 1896-1908
- Relaterad länk:
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https://doi.org/10.1...
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https://uu.diva-port... (primary) (Raw object)
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https://urn.kb.se/re...
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https://doi.org/10.1...
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http://kipublication...
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Abstract
Ämnesord
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- The tetracycline repressor (tetR)-regulated system is a widely used tool to specifically control gene expression in mammalian cells. Based on this system, we generated a human osteosarcoma cell line, which allows for the inducible expression of an EGFP fusion of the TAR DNA-binding protein 43 (TDP-43), which has been linked to neurodegenerative diseases. Consistent with previous findings, TDP-43 overexpression led to the accumulation of aggregates and limited the viability of U2OS. Using this inducible system, we conducted a chemical screen with a library that included FDA-approved drugs. While the primary screen identified several compounds that prevented TDP-43 toxicity, further experiments revealed that these chemicals abrogated the doxycycline-dependent TDP-43 expression. This antagonistic effect was observed with both doxycycline and tetracycline, and in several Tet-On cell lines expressing different genes, confirming the general effect of these compounds as inhibitors of the tetR system. Using the same cell line, a genome-wide CRISPR/Cas9 screen identified epigenetic regulators such as the G9a methyltransferase and TRIM28 as potential modifiers of TDP-43 toxicity. Yet again, further experiments revealed that G9a inhibition or TRIM28 loss prevented doxycycline-dependent expression of TDP-43. In summary, we have identified new chemical and genetic regulators of the tetR system, thereby raising awareness of the limitations of this approach to conduct chemical or genetic screening in mammalian cells.
Ämnesord
- MEDICIN OCH HÄLSOVETENSKAP -- Medicinsk bioteknologi -- Biomedicinsk laboratorievetenskap/teknologi (hsv//swe)
- MEDICAL AND HEALTH SCIENCES -- Medical Biotechnology -- Biomedical Laboratory Science/Technology (hsv//eng)
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