RRC ID 63909
Author Yesbolatova A, Saito Y, Kitamoto N, Makino-Itou H, Ajima R, Nakano R, Nakaoka H, Fukui K, Gamo K, Tominari Y, Takeuchi H, Saga Y, Hayashi KI, Kanemaki MT.
Title The auxin-inducible degron 2 technology provides sharp degradation control in yeast, mammalian cells, and mice.
Journal Nat Commun
Abstract Protein knockdown using the auxin-inducible degron (AID) technology is useful to study protein function in living cells because it induces rapid depletion, which makes it possible to observe an immediate phenotype. However, the current AID system has two major drawbacks: leaky degradation and the requirement for a high dose of auxin. These negative features make it difficult to control precisely the expression level of a protein of interest in living cells and to apply this method to mice. Here, we overcome these problems by taking advantage of a bump-and-hole approach to establish the AID version 2 (AID2) system. AID2, which employs an OsTIR1(F74G) mutant and a ligand, 5-Ph-IAA, shows no detectable leaky degradation, requires a 670-times lower ligand concentration, and achieves even quicker degradation than the conventional AID. We demonstrate successful generation of human cell mutants for genes that were previously difficult to deal with, and show that AID2 achieves rapid target depletion not only in yeast and mammalian cells, but also in mice.
Volume 11(1)
Pages 5701
Published 2020-11-11
DOI 10.1038/s41467-020-19532-z
PII 10.1038/s41467-020-19532-z
PMID 33177522
IF 12.121
Resource
DNA material pAY7 (RDB18333) pAY15 (RDB18334) pAY19 (RDB18335) pMK381 (RDB18337) pAAV-hSyn-OsTIR1(WT) (RDB18338) pAAV-hSyn-OsTIR1(F74G) (RDB18339) pAAV-hSyn-OsTIR1(F74A) (RDB18340) pMK198 (RDB18341) pMK419 (RDB18342) pMK425 (RDB18343) pMK262 (RDB18344) pMK265 (RDB18345) RAD21 C-tag CIRPSR (RDB18346) pMK233 (RDB18347) pMK296 (RDB18348) pMK297 (RDB18349) DHC1-mIAA7 Neo (RDB18350) DHC1-mIAA7 Hygro (RDB18351) DHC1 CRISPR (RDB18352) CTCF-mAC donor (Neo) (RDB18353) CTCF-mAC donor (Hygro) (RDB18354) CTCF-C CRISPR (RDB18355) SMC2-mAC donor (Hygro) (RDB18356) SMC2 CRISPR (RDB18357) mAID-BRD4 donor (RDB18358) BRD4-N CRISPR (RDB18359) pMK321 (RDB18360) pMK322 (RDB18361) pMK312 (RDB18362) pMK427 (RDB18363) pMK411 (RDB18364)