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Open Access Highly Accessed Research article

alpha-Sarcin catalytic activity is not required for cytotoxicity

Spencer C Alford12*, Joel D Pearson4, Amanda Carette2, Robert J Ingham4 and Perry L Howard13

Author Affiliations

1 Centre for Biomedical Research, University of Victoria, PO Box 3020 Station CSC Victoria, British Columbia, V8W 3N5, Canada

2 Department of Biochemistry and Microbiology, University of Victoria, PO Box 3055 Station CSC Victoria, British Columbia, V8W 3P6, Canada

3 Department of Biology, University of Victoria PO Box 3020 Station CSC Victoria, British Columbia, V8W 3N5, Canada

4 Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, T6G 2H7, Canada

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BMC Biochemistry 2009, 10:9  doi:10.1186/1471-2091-10-9

Published: 3 April 2009

Abstract

Background

α-Sarcin is a protein toxin produced by Aspergillus giganteus. It belongs to a family of cytotoxic ribonucleases that inactivate the ribosome and inhibit protein synthesis. α-Sarcin cleaves a single phosphodiester bond within the RNA backbone of the large ribosomal subunit, which makes the ribosome unrecognizable to elongation factors and, in turn, blocks protein synthesis. Although it is widely held that the protein synthesis inhibition caused by the toxin leads to cell death, it has not been directly shown that catalytically inactive mutants of α-sarcin are non-toxic when expressed directly within the cytoplasm of cells. This is important since recent studies have cast doubt on whether protein synthesis inhibition is sufficient to initiate apoptosis.

Results

In this report, we assay α-sarcin cytotoxicity and ability to inhibit protein synthesis by direct cytoplasmic expression. We show that mutations in α-sarcin, which impair α-sarcin's ability to inhibit protein synthesis, do not affect its cytotoxicity. The mutants are unable to activate JNK, confirming that the sarcin-ricin loop remains intact and that the α-sarcin mutants are catalytically inactive. In addition, both mutant and wildtype variants of α-sarcin localize to the nucleus and cytoplasm, where they co-localize with ribosomal marker RPS6.

Conclusion

We conclude that although protein synthesis inhibition likely contributes to cell death, it is not required. Thus, our results suggest that α-sarcin can promote cell death through a previously unappreciated mechanism that is independent of rRNA cleavage and JNK activation.