Methods for measuring HMGB1 release during immunogenic cell death.
Antineoplastic Agents
/ pharmacology
Cell Line, Tumor
Cell Membrane
/ drug effects
Cell Nucleus
/ drug effects
Drug Discovery
/ instrumentation
Green Fluorescent Proteins
/ genetics
HMGB1 Protein
/ analysis
High-Throughput Screening Assays
/ instrumentation
Humans
Immunity, Innate
/ drug effects
Immunogenic Cell Death
/ drug effects
Molecular Probes
/ genetics
Neoplasms
/ drug therapy
Nuclear Localization Signals
/ genetics
Recombinant Fusion Proteins
/ genetics
Signal Transduction
/ drug effects
Streptavidin
/ genetics
Anthracyclines
Cancer
High-throughput screening
Immunogenic cell death (ICD)
Retention using selective hooks (RUSH) assay
Journal
Methods in enzymology
ISSN: 1557-7988
Titre abrégé: Methods Enzymol
Pays: United States
ID NLM: 0212271
Informations de publication
Date de publication:
2019
2019
Historique:
entrez:
16
11
2019
pubmed:
16
11
2019
medline:
2
6
2020
Statut:
ppublish
Résumé
The exodus of the alarmin high mobility group box 1 (HMGB1) from the nucleus constitutes a crucial cellular danger signal and manifests as a sequential process in which HMGB1 first exits the nucleus into the cytoplasm and then is secreted or passively released through the permeabilized plasma membrane. Extracellular HMGB1 can interact with pattern recognition receptors to stimulate innate immune responses. Here, we describe a discovery pipeline for the identification of pharmacological agents endowed with HMGB1 releasing properties. The "retention using selective hooks" (RUSH) system in which a streptavidin-NLS3 fusion protein serves as a nuclear hook to sequester streptavidin-binding peptide (SBP) fused with HMGB1 and green fluorescent protein (GFP) allowed for synchronizing HMGB1 increase. Thus, exclusively in the presence of biotin, which liberates HMGB1-SBP-GFP from its nuclear hook, immunogenic cell death (ICD) inducers such as anthracyclines are able to cause the nucleo-cytoplasmic translocation of HMGB1-SBP-GFP. This system facilitates the identification of HMGB1 releasing agents in medium- to high-throughput screening assays.
Identifiants
pubmed: 31727240
pii: S0076-6879(19)30149-1
doi: 10.1016/bs.mie.2019.05.001
pii:
doi:
Substances chimiques
Antineoplastic Agents
0
HMGB1 Protein
0
HMGB1 protein, human
0
Molecular Probes
0
Nuclear Localization Signals
0
Recombinant Fusion Proteins
0
Green Fluorescent Proteins
147336-22-9
Streptavidin
9013-20-1
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
177-193Informations de copyright
© 2019 Elsevier Inc. All rights reserved.