Yeast Reporter Assay to Identify Cellular Components of Ricin Toxin A Chain Trafficking
AbstractRTA, the catalytic A-subunit of the ribosome inactivating A/B toxin ricin, inhibits eukaryotic protein biosynthesis by depurination of 28S rRNA. Although cell surface binding of ricin holotoxin is mainly mediated through its B-subunit (RTB), sole application of RTA is also toxic, albeit to a significantly lower extent, suggesting alternative pathways for toxin uptake and transport. Since ricin toxin trafficking in mammalian cells is still not fully understood, we developed a GFP-based reporter assay in yeast that allows rapid identification of cellular components required for RTA uptake and subsequent transport through a target cell. We hereby show that Ypt6p, Sft2p and GARP-complex components play an important role in RTA transport, while neither the retromer complex nor COPIB vesicles are part of the transport machinery. Analyses of yeast knock-out mutants with chromosomal deletion in genes whose products regulate ADP-ribosylation factor GTPases (Arf-GTPases) and/or retrograde Golgi-to-ER (endoplasmic reticulum) transport identified Sso1p, Snc1p, Rer1p, Sec22p, Erv46p, Gea1p and Glo3p as novel components in RTA transport, suggesting the developed reporter assay as a powerful tool to dissect the multistep processes of host cell intoxication in yeast. View Full-Text
Share & Cite This Article
Becker, B.; Schnöder, T.; Schmitt, M.J. Yeast Reporter Assay to Identify Cellular Components of Ricin Toxin A Chain Trafficking. Toxins 2016, 8, 366.
Becker B, Schnöder T, Schmitt MJ. Yeast Reporter Assay to Identify Cellular Components of Ricin Toxin A Chain Trafficking. Toxins. 2016; 8(12):366.Chicago/Turabian Style
Becker, Björn; Schnöder, Tina; Schmitt, Manfred J. 2016. "Yeast Reporter Assay to Identify Cellular Components of Ricin Toxin A Chain Trafficking." Toxins 8, no. 12: 366.