Eeyarestatin 1 interferes with both retrograde and anterograde intracellular trafficking pathways.

Abstract

BACKGROUND:

The small molecule Eeyarestatin I (ESI) inhibits the endoplasmic reticulum (ER)-cytosol dislocation and subsequent degradation of ERAD (ER associated protein degradation) substrates. Toxins such as ricin and Shiga/Shiga-like toxins (SLTx) are endocytosed and trafficked to the ER. Their catalytic subunits are thought to utilise ERAD-like mechanisms to dislocate from the ER into the cytosol, where a proportion uncouples from the ERAD process, recovers a catalytic conformation and destroys their cellular targets. We therefore investigated ESI as a potential inhibitor of toxin dislocation.

METHODOLOGY AND PRINCIPAL FINDINGS:

Using cytotoxicity measurements, we found no role for ES(I) as an inhibitor of toxin dislocation from the ER, but instead found that for SLTx, ESI treatment of cells was protective by reducing the rate of toxin delivery to the ER. Microscopy of the trafficking of labelled SLTx and its B chain (lacking the toxic A chain) showed a delay in its accumulation at a peri-nuclear location, confirmed to be the Golgi by examination of SLTx B chain metabolically labelled in the trans-Golgi cisternae. The drug also reduced the rate of endosomal trafficking of diphtheria toxin, which enters the cytosol from acidified endosomes, and delayed the Golgi-specific glycan modifications and eventual plasma membrane appearance of tsO45 VSV-G protein, a classical marker for anterograde trafficking.

CONCLUSIONS AND SIGNIFICANCE:

ESI acts on one or more components that function during vesicular transport, whilst at least one retrograde trafficking pathway, that of ricin, remains unperturbed.

• Authors: Mina-Olga Aletrari, Craig McKibbin, Helen Williams, Vidya Pawar, Paola Pietroni, J Michael Lord, Sabine L Flitsch, Roger Whitehead, Eileithyia Swanton, Stephen High, Robert A Spooner
• Journal: PloS one (PLoS One) Vol. 6 Issue 7 Pg. e22713 ( 2011) ISSN: 1932-6203 [Electronic] United States
• PMID: 21799938 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• 1-(4-chlorophenyl)-3-(3-(4-chlorophenyl)-5,5-dimethyl-1-(3-(5-nitrofuran-2-yl)allyldienehydrazinocarbonylmethyl)-2-oxoimidazolidin-4-yl)-1-hydroxyurea
• Diphtheria Toxin
• G protein, vesicular stomatitis virus
• Hydrazones
• Membrane Glycoproteins
• Viral Envelope Proteins
• Shiga Toxin
• Ricin
• Hydroxyurea

Topics

• Biological Transport (drug effects)
• Cytosol (drug effects, metabolism)
• Diphtheria Toxin (metabolism, toxicity)
• Endoplasmic Reticulum (drug effects, metabolism)
• HeLa Cells
• Humans
• Hydrazones (pharmacology)
• Hydroxyurea (analogs & derivatives, pharmacology)
• Intracellular Membranes (drug effects, metabolism)
• Intracellular Space (drug effects, metabolism)
• Membrane Glycoproteins (metabolism)
• Ricin (metabolism, toxicity)
• Shiga Toxin (metabolism, toxicity)
• Time Factors
• Viral Envelope Proteins (metabolism)