Retro-2 protects cells from ricin toxicity by inhibiting ASNA1-mediated ER targeting and insertion of tail-anchored proteins.
CRISPR
Retro-2
TRC pathway
cell biology
genetics
genomics
human
Journal
eLife
ISSN: 2050-084X
Titre abrégé: Elife
Pays: England
ID NLM: 101579614
Informations de publication
Date de publication:
01 11 2019
01 11 2019
Historique:
received:
13
05
2019
accepted:
28
10
2019
pubmed:
2
11
2019
medline:
28
5
2020
entrez:
2
11
2019
Statut:
epublish
Résumé
The small molecule Retro-2 prevents ricin toxicity through a poorly-defined mechanism of action (MOA), which involves halting retrograde vesicle transport to the endoplasmic reticulum (ER). CRISPRi genetic interaction analysis revealed Retro-2 activity resembles disruption of the transmembrane domain recognition complex (TRC) pathway, which mediates post-translational ER-targeting and insertion of tail-anchored (TA) proteins, including SNAREs required for retrograde transport. Cell-based and in vitro assays show that Retro-2 blocks delivery of newly-synthesized TA-proteins to the ER-targeting factor ASNA1 (TRC40). An ASNA1 point mutant identified using CRISPR-mediated mutagenesis abolishes both the cytoprotective effect of Retro-2 against ricin and its inhibitory effect on ASNA1-mediated ER-targeting. Together, our work explains how Retro-2 prevents retrograde trafficking of toxins by inhibiting TA-protein targeting, describes a general CRISPR strategy for predicting the MOA of small molecules, and paves the way for drugging the TRC pathway to treat broad classes of viruses known to be inhibited by Retro-2.
Identifiants
pubmed: 31674906
doi: 10.7554/eLife.48434
pii: 48434
pmc: PMC6858068
doi:
pii:
Substances chimiques
2-(((5-methyl-2-thienyl)methylene)amino)-N-phenylbenzamide
0
Benzamides
0
GET3 protein, human
0
Membrane Proteins
0
Thiophenes
0
Ricin
9009-86-3
Arsenite Transporting ATPases
EC 3.6.3.16
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Langues
eng
Sous-ensembles de citation
IM
Subventions
Organisme : NCI NIH HHS
ID : T32 CA009302
Pays : United States
Organisme : NICHD NIH HHS
ID : DP2 HD084069
Pays : United States
Organisme : NIH HHS
ID : DP2HD084069
Pays : United States
Organisme : NIGMS NIH HHS
ID : R35 GM127136
Pays : United States
Organisme : NIGMS NIH HHS
ID : 1R35GM127136
Pays : United States
Organisme : NHGRI NIH HHS
ID : T32 HG000044
Pays : United States
Organisme : NLM NIH HHS
ID : T32 LM012409
Pays : United States
Organisme : NIH HHS
ID : 1DP2HD084069-01
Pays : United States
Organisme : National Science Foundation
ID : DGE-1656518
Pays : International
Organisme : NIH HHS
ID : T32 HG000044
Pays : United States
Organisme : National Science Foundation
ID : DGE-114747
Pays : International
Informations de copyright
© 2019, Morgens et al.
Déclaration de conflit d'intérêts
DM, CC, AK, NW, AL, MD, CT, GH, AL, KH, NP, JZ, EH, PA, AD, DY, RA, JS, VD, MB No competing interests declared
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