Disruption of iron homeostasis increases phosphine toxicity in caenorhabditis elegans

The aim of this study is to identify the biochemical mechanism of phosphine toxicity and resistance, using Caenorhabditis elegans as a model organism. To date, the precise mode of phosphine action is unclear. In this report, we demonstrate the following dose-dependent actions of phosphine, in vitro: (1) reduction of ferric iron (Fe3+) to ferrous iron (Fe2+), (2) release of iron from horse ferritin, (3) and the peroxidation of lipid as a result of iron release from ferritin. Using in situ hybridization, we show that the ferritin genes of C. elegans, both ferritin-1 and ferritin-2, are expressed along the digestive tract with greatest expression at the proximal and distal ends. Basal expression of the ferritin-2 gene, as determined by quantitative PCR, is approximately 80 times that of ferritin-1. However, transcript levels of ferritin-1 are induced at least 20-fold in response to phosphine, whereas there is no change in the level of ferritin-2. This resembles the reported pattern of ferritin gene regulation by iron, suggesting that phosphine toxicity may be related to an increase in the level of free iron. Indeed, iron overload increases phosphine toxicity in C. elegans at least threefold. Moreover, we demonstrate that suppression of ferritin-2 gene expression by RNAi, significantly increases sensitivity to phosphine. This study identifies similarities between phosphine toxicity and iron overload and demonstrates that phosphine can trigger iron release from storage proteins, increasing lipid peroxidation, leading to cell injury and/or cell death.

Differing mechanisms of simple nitrile formation on glucosinolate degradation in Lepidium sativum and Nasturtium officinale seeds.

Glucosinolates are sulphur-containing glycosides found in brassicaceous plants that can be hydrolysed enzymatically by plant myrosinase or non-enzymatically to form primarily isothiocyanates and/or simple nitriles. From a human health perspective, isothiocyanates are quite important because they are major inducers of carcinogen-detoxifying enzymes. Two of the most potent inducers are benzyl isothiocyanate (BITC) present in garden cress (Lepidium sativum), and phenylethyl isothiocyanate (PEITC) present in watercress (Nasturtium officinale). Previous studies on these salad crops have indicated that significant amounts of simple nitriles are produced at the expense of the isothiocyanates. These studies also suggested that nitrile formation may occur by different pathways: (1) under the control of specifier protein in garden cress and (2) by an unspecified, non-enzymatic path in watercress. In an effort to understand more about the mechanisms involved in simple nitrile formation in these species, we analysed their seeds for specifier protein and myrosinase activities, endogenous iron content and glucosinolate degradation products after addition of different iron species, specific chelators and various heat treatments. We confirmed that simple nitrile formation was predominantly under specifier protein control (thiocyanate-forming protein) in garden cress seeds. Limited thermal degradation of the major glucosinolate, glucotropaeolin (benzyl glucosinolate), occurred when seed material was heated to >120 degrees C. In the watercress seeds, however, we show for the first time that gluconasturtiin (phenylethyl glucosinolate) undergoes a non-enzymatic, iron-dependent degradation to a simple nitrile. On heating the seeds to 120 degrees C or greater, thermal degradation of this heat-labile glucosinolate increased simple nitrile levels many fold.