Trichothecene mycotoxins synthesized by Fusarium species are potent inhibitors of eukaryotic translation. They are encountered in both the environment and in food, posing a threat to human and animal health. They have diverse roles in the cell that are not limited to the inhibition of protein synthesis. To understand the trichothecene mechanism of action, we screened the yeast knockout library to identify genes whose deletion confers resistance to trichothecin (Tcin). The largest group of resistant strains affected mitochondrial function, suggesting a role for fully active mitochondria in trichothecene toxicity. Tcin inhibited mitochondrial translation in the wild-type strain to a greater extent than in the most resistant strains, implicating mitochondrial translation as a previously unrecognized site of action. The Tcin-resistant strains were cross-resistant to anisomycin and chloramphenicol, suggesting that Tcin targets the peptidyltransferase center of mitochondrial ribosomes. Tcin-induced cell death was partially rescued by mutants that regulate mitochondrial fusion and maintenance of the tubular morphology of mitochondria. Treatment of yeast cells with Tcin led to the fragmentation of the tubular mitochondrial network, supporting a role for Tcin in disruption of mitochondrial membrane morphology. These results provide genome-wide insight into the mode of action of trichothecene mycotoxins and uncover a critical role for mitochondrial translation and membrane maintenance in their toxicity.Fusarium head blight ͉ ribosome ͉ translation ͉ deoxynivalenol ͉ mycotoxin

Ricin A chain (RTA) inhibits protein synthesis by removing a specific adenine from the highly conserved ␣-sarcin/ricin loop in the large rRNA. Expression of RTA with its own signal sequence in yeast resulted in its translocation into the endoplasmic reticulum (ER) and subsequent glycosylation. Because RTA must unfold within the ER, it may be vulnerable to host defenses, such as the unfolded protein response (UPR). UPR was induced in cells expressing an active site mutant but not the wild type RTA, indicating that the active site of RTA played a role in perturbing the ER stress response. The inactive RTA without the signal sequence did not induce UPR, indicating that translocation into the ER was critical for induction of UPR. The wild type RTA inhibited activation of UPR not only due to ER stress induced by the protein itself but also by global effectors such as tunicamycin and dithiothreitol. Mature RTA without the signal sequence also inhibited UPR, providing evidence that inhibition of UPR occurred on the cytosolic face of the ER. RTA could not inhibit UPR when the spliced form of HAC1 mRNA was provided in trans, indicating that it had a direct effect on UPR upstream of HAC1-dependent transcriptional activation. Only the precursor form of HAC1 mRNA was detected in cells expressing RTA after exposure to ER stress, demonstrating that ricin inhibits activation of UPR by preventing HAC1 mRNA splicing. The RTA mutants that depurinated ribosomes but did not kill cells were not able to inhibit activation of UPR by tunicamycin, providing evidence that the inability to activate UPR in response to ER stress contributes to the cytotoxicity of ricin.

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Aims: Inactivation processes using heat are widely used for disinfection and sterilization. Dry heat sterilization of spacecraft equipment has been the preferred microbial inactivation method as part of interplanetary travel protection strategies. An antimicrobial model, based on temperature and exposure time based on experimental data, was developed to provide reliable sterilization processes to be used for interplanetary applications.Methods and Results: Bacillus atrophaeus spores, traditionally used to challenge dry heat sterilization processes, were tested over a range of temperatures in comparison with spores of Bacillus canaveralius that have been shown to have a higher heat resistance profile. D-value and Z-values were determined and used to develop a mathematical model for parametric sterilization applications. The impact of the presence of a contaminating soil, representative of Mars dust, was also tested to verify the practical application of the model to reduce the risk of microbial contamination in such environments. Conclusion:The sterilization model developed can be used as an intrinsic part of risk reduction strategies for interplanetary protection.Significance and Impact: Forward and backward planetary protection strategies to reduce the risks of microbial contamination during interplanetary exploration and research is an important consideration. The development of a modern sterilization model, with consideration of microorganisms identified with higher levels of heat resistance than traditionally deployed in terrestrial applications, allows for the consideration of optimal inactivation processes to define minimum criteria for engineering design. The ability to inactivate living microorganisms, as well as to degrade biomolecules, provides a reliable method to reduce the risk of known and potentially unknown contaminants in future applications.