Development of AlissAID system targeting GFP or mCherry fusion protein

Yoshitaka Ogawa; Kohei Nishimura; Keisuke Obara; Takumi Kamura

doi:10.1371/journal.pgen.1010731

Development of AlissAID system targeting GFP or mCherry fusion protein

PLoS Genet. 2023 Jun 14;19(6):e1010731. doi: 10.1371/journal.pgen.1010731. eCollection 2023 Jun.

Authors

Yoshitaka Ogawa¹, Kohei Nishimura¹, Keisuke Obara¹, Takumi Kamura¹

Affiliation

¹ Department of Biological Science, Division of Natural Science, Graduate School of Science, Nagoya University, Nagoya, Japan.

Abstract

Conditional control of target proteins using the auxin-inducible degron (AID) system provides a powerful tool for investigating protein function in eukaryotes. Here, we established an Affinity-linker based super-sensitive auxin-inducible degron (AlissAID) system in budding yeast by using a single domain antibody (a nanobody). In this system, target proteins fused with GFP or mCherry were degraded depending on a synthetic auxin, 5-Adamantyl-IAA (5-Ad-IAA). In AlissAID system, nanomolar concentration of 5-Ad-IAA induces target degradation, thus minimizing the side effects from chemical compounds. In addition, in AlissAID system, we observed few basal degradations which was observed in other AID systems including ssAID system. Furthermore, AlissAID based conditional knockdown cell lines are easily generated by using budding yeast GFP Clone Collection. Target protein, which has antigen recognition sites exposed in cytosol or nucleus, can be degraded by the AlissAID system. From these advantages, the AlissAID system would be an ideal protein-knockdown system in budding yeast cells.

Copyright: © 2023 Ogawa et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cell Line
Cell Nucleus
Cytosol
Drug-Related Side Effects and Adverse Reactions*
Indoleacetic Acids
Saccharomycetales*

Substances

Indoleacetic Acids

Grants and funding

KN received support from the Japan Society for the Promotion of Science KAKENHI Grant Numbers, JP19K06611, JP20K21423 and JP22K05558. KN also received support from Kato Memorial Bioscience Foundation and Institute for Fermentation, Osaka. KO received support from the Japan Society for the Promotion of Science KAKENHI Grant Number, JP22K06141 and from Institute for Fermentation, Osaka. TK received support from the Japan Society for the Promotion of Science KAKENHI Grant Number, JP20H03208. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.