Linked Life Data

20659495

Source:http://linkedlifedata.com/resource/pubmed/id/20659495

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
6
pubmed:dateCreated
2010-8-30
pubmed:abstractText
Pseudomonas exotoxin A (PE) is cytotoxic for eukaryotic cells because it enters cells by receptor-mediated endocytosis, translocates to the cell cytosol and ADP-ribosylates elongation factor 2 (EF2). However, the interaction of this toxin with eukaryotic cells and the mechanism of PE-mediated cell death have not been extensively characterized. The feasibility of carrying out a genome-wide RNAi screen, makes Drosophila melanogaster S2 cells as a good model system to identify essential genes in PE-mediated cytotoxicity, provided a suitable multi-well assay is developed. Here, using the alamarBlue viability assay, we show that Drosophila S2 cells are sensitive to PE at picomolar concentrations and that toxin treatments provoke an increase in caspase activity. This prompted us to use RNAi to characterize the mechanism of cell death. Results indicated that PE-mediated death of S2 cells was dependent on the presence of diphthamide, the post translational modification of EF2, and on the presence of Drice, the terminal caspase of insect cells. RNAi to drice or chemical inhibition of caspase action by z-VAD-fmk protected cells from PE-mediated death. Protection from death by RNAi or z-VAD-fmk did not interfere with toxin delivery to the cytosol leading to inhibition of protein synthesis. Using a convenient alamarBlue assay, our data confirms the cytotoxicity of PE for S2 cells and establishes apoptosis as the mode of PE-mediated death. This confirms the suitability of Drosophila cells as a convenient and simple model to elucidate the role of specific genes and proteins required for PE action.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
http://linkedlifedata.com/resource/pubmed/chemical/ADP Ribose Transferases, http://linkedlifedata.com/resource/pubmed/chemical/Amino Acid Chloromethyl Ketones, http://linkedlifedata.com/resource/pubmed/chemical/Bacterial Toxins, http://linkedlifedata.com/resource/pubmed/chemical/Caspases, http://linkedlifedata.com/resource/pubmed/chemical/Drosophila Proteins, http://linkedlifedata.com/resource/pubmed/chemical/Exotoxins, http://linkedlifedata.com/resource/pubmed/chemical/Histidine, http://linkedlifedata.com/resource/pubmed/chemical/Ice protein, Drosophila, http://linkedlifedata.com/resource/pubmed/chemical/RNA, Small Interfering, http://linkedlifedata.com/resource/pubmed/chemical/Virulence Factors, http://linkedlifedata.com/resource/pubmed/chemical/benzyloxycarbonylvalyl-alanyl-aspart..., http://linkedlifedata.com/resource/pubmed/chemical/diphthamide, http://linkedlifedata.com/resource/pubmed/chemical/toxA protein, Pseudomonas aeruginosa
pubmed:status
MEDLINE
pubmed:month
Nov
pubmed:issn
1879-3150
pubmed:author
pubmed:copyrightInfo
(c) 2010 Elsevier Ltd. All rights reserved.
pubmed:issnType
Electronic
pubmed:volume
56
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
1025-34
pubmed:meshHeading
pubmed:year
2010
pubmed:articleTitle
Pseudomonas exotoxin kills Drosophila S2 cells via apoptosis.
pubmed:affiliation
Biotherapy Section, Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, NIH, HHS, 37 Convent Dr, Bethesda, MD 20892, USA.
pubmed:publicationType
Journal Article