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Ribosome Depurination Is Not Sufficient for Ricin-Mediated Cell Death in Saccharomyces cerevisiae ^,

Xiao-Ping Li,^1,3, Marianne Baricevic,^1,2,3, Hemalatha Saidasan,^1,3 and Nilgun E. Tumer^1,2,3*

Biotechnology Center for Agriculture and the Environment,¹ Graduate Program in Molecular Biosciences,² Department of Plant Biology and Pathology, Cook College, Rutgers University, New Brunswick, New Jersey 08901-8520³

Received 11 August 2006/ Returned for modification 22 September 2006/ Accepted 13 October 2006

The plant toxin ricin is one of the most potent and lethal substances known. Ricin inhibits protein synthesis by removing a specific adenine from the highly conserved -sarcin/ricin loop in the large rRNA. Very little is known about how ricin interacts with ribosomes and the molecular mechanism by which it kills cells. To gain insight to the mechanism of ricin-induced cell death, we set up yeast (Saccharomyces cerevisiae) as a simple and genetically tractable system to isolate mutants defective in cytotoxicity. Ribosomes were depurinated in yeast cells expressing the precursor form of the A chain of ricin (pre-RTA), and these cells displayed apoptotic markers such as nuclear fragmentation, chromatin condensation, and accumulation of reactive oxygen species. We conducted a large-scale mutagenesis of pre-RTA and isolated a panel of nontoxic RTA mutants based on their inability to kill yeast cells. Several nontoxic RTA mutants depurinated ribosomes and inhibited translation to the same extent as wild-type RTA in vivo. The mutant proteins isolated from yeast depurinated ribosomes in vitro, indicating that they were catalytically active. However, cells expressing these mutants did not display hallmarks of apoptosis. These results provide the first evidence that the ability to depurinate ribosomes and inhibit translation does not always correlate with ricin-mediated cell death, indicating that ribosome depurination and translation inhibition do not account entirely for the cytotoxicity of ricin.

Published ahead of print on 13 November 2006.

Editor: A. D. O'Brien

Supplemental material for this article may be found at http://iai.asm.org/.

The first two authors should be regarded as joint first authors.

This article has been cited by other articles:

• Parikh, B. A., Tortora, A., Li, X.-P., Tumer, N. E. (2008). Ricin Inhibits Activation of the Unfolded Protein Response by Preventing Splicing of the HAC1 mRNA. J. Biol. Chem. 283: 6145-6153 [Abstract] [Full Text]