Genome-wide conditional degron libraries for functional genomics.
| Autor: | Gameiro E; Institute of Molecular Biology , Mainz, Germany., Juárez-Núñez KA; Institute of Molecular Biology , Mainz, Germany., Fung JJ; Institute of Molecular Biology , Mainz, Germany., Shankar S; Institute of Molecular Biology , Mainz, Germany., Luke B; Institute of Molecular Biology , Mainz, Germany.; Johannes Gutenberg University Mainz, Institute for Developmental Neurology , Mainz, Germany., Khmelinskii A; Institute of Molecular Biology , Mainz, Germany.; Institute for Quantitative and Computational Biosciences, Johannes Gutenberg University Mainz , Mainz, Germany. |
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| Jazyk: | angličtina |
| Zdroj: | The Journal of cell biology [J Cell Biol] 2025 Feb 03; Vol. 224 (2). Date of Electronic Publication: 2024 Dec 18. |
| DOI: | 10.1083/jcb.202409007 |
| Abstrakt: | Functional genomics with libraries of knockout alleles is limited to non-essential genes and convoluted by the potential accumulation of suppressor mutations in knockout backgrounds, which can lead to erroneous functional annotations. To address these limitations, we constructed genome-wide libraries of conditional alleles based on the auxin-inducible degron (AID) system for inducible degradation of AID-tagged proteins in the budding yeast Saccharomyces cerevisiae. First, we determined that N-terminal tagging is at least twice as likely to inadvertently impair protein function across the proteome. We thus constructed two libraries with over 5,600 essential and non-essential proteins fused at the C-terminus with an AID tag and an optional fluorescent protein. Approximately 90% of AID-tagged proteins were degraded in the presence of the auxin analog 5-Ph-IAA, with initial protein abundance and tag accessibility as limiting factors. Genome-wide screens for DNA damage response factors revealed a role for the glucose signaling factor GSF2 in resistance to hydroxyurea, highlighting how the AID libraries extend the yeast genetics toolbox. (© 2024 Gameiro et al.) |
| Databáze: | MEDLINE |
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