Creation and validation of a proteome-wide yeast library for protein detection and analysis

Din Baruch; Ioannis Tsirkas; Ehud Sass; Benjamin Dubreuil; Yeynit Asraf; Amir Aharoni; Maya Schuldiner; Ofir Klein

doi:10.1242/jcs.263848

Creation and validation of a proteome-wide yeast library for protein detection and analysis

Din Baruch
, Ioannis Tsirkas
, Ehud Sass
, Benjamin Dubreuil
, Yeynit Asraf
, Amir Aharoni
, Maya Schuldiner^*
, Ofir Klein^*

^*Corresponding author for this work

Department of Molecular Genetics

Research output: Contribution to journal › Article › peer-review

2 Citations (Scopus)

42 Downloads (Pure)

Abstract

A significant challenge in cell biology is to uncover the function of uncharacterized proteins. Surprisingly, a quarter of the proteome is still poorly understood even in the most well-studied model organisms. Systematic methodologies, including the use of tagged protein collections, have emerged as a powerful approach to address this gap. Despite the availability of proteome-wide collections featuring various fused proteins, the impact of different tags on protein function highlights the need for diversifying the tags used for functional genomic studies. To rise to this challenge, we created a proteome-wide collection of yeast strains in which proteins are N-terminally tagged with the broadly utilized and compact hemagglutinin (HA) epitope. We showcase the potential uses of our library for systematically evaluating protein size, abundance and localization using an in vivo labeling approach. Our characterization underscores the potential utility of a proteome-wide HA-tagged library in revealing novel aspects of cell biology, providing an additional powerful tool for functional genomics.

Original language	English
Article number	jcs263848
Number of pages	10
Journal	Journal of Cell Science
Volume	138
Issue number	16
DOIs	https://doi.org/10.1242/jcs.263848
Publication status	Published - Aug 2025

Funding

We thank Rosario Valenti and Olga Beresh for critical reading of the manuscript and helpful suggestions. We thank Prof. Michael Toledano for the idea to create this library. We thank Dr Yoav Peleg for help with plasmid generation. The robotic system in the Schuldiner laboratory was purchased through the kind support of the Blythe Brenden-Mann Foundation. M.S. is Incumbent of the Dr Gilbert Omenn and Martha Darling Professorial Chair in Molecular Genetics. This work was supported by a Chan Zuckerberg Initiative (CZI) grant (2023-331952) and by the European Union European Research Council (ERC) CoG (OnTarget 864068) as well as by the Institute for Environmental Sustainability (IES) at the Weizmann Institute of Science. Open Access funding provided by the Weizmann Institute of Science. Deposited in PMC for immediate release.

All Science Journal Classification (ASJC) codes

Cell Biology

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ms_JCellSci_CreationandVadidationofProteome_PV2025Final published version, 3.81 MBLicence: CC BY

Cite this

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abstract = "A significant challenge in cell biology is to uncover the function of uncharacterized proteins. Surprisingly, a quarter of the proteome is still poorly understood even in the most well-studied model organisms. Systematic methodologies, including the use of tagged protein collections, have emerged as a powerful approach to address this gap. Despite the availability of proteome-wide collections featuring various fused proteins, the impact of different tags on protein function highlights the need for diversifying the tags used for functional genomic studies. To rise to this challenge, we created a proteome-wide collection of yeast strains in which proteins are N-terminally tagged with the broadly utilized and compact hemagglutinin (HA) epitope. We showcase the potential uses of our library for systematically evaluating protein size, abundance and localization using an in vivo labeling approach. Our characterization underscores the potential utility of a proteome-wide HA-tagged library in revealing novel aspects of cell biology, providing an additional powerful tool for functional genomics.",

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AU - Aharoni, Amir

AU - Schuldiner, Maya

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AB - A significant challenge in cell biology is to uncover the function of uncharacterized proteins. Surprisingly, a quarter of the proteome is still poorly understood even in the most well-studied model organisms. Systematic methodologies, including the use of tagged protein collections, have emerged as a powerful approach to address this gap. Despite the availability of proteome-wide collections featuring various fused proteins, the impact of different tags on protein function highlights the need for diversifying the tags used for functional genomic studies. To rise to this challenge, we created a proteome-wide collection of yeast strains in which proteins are N-terminally tagged with the broadly utilized and compact hemagglutinin (HA) epitope. We showcase the potential uses of our library for systematically evaluating protein size, abundance and localization using an in vivo labeling approach. Our characterization underscores the potential utility of a proteome-wide HA-tagged library in revealing novel aspects of cell biology, providing an additional powerful tool for functional genomics.

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Creation and validation of a proteome-wide yeast library for protein detection and analysis

Abstract

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