Abstract
All terrestrial organisms are subject to evolutionary pressures associated with natural sources of ionizing radiation (IR). The legacy of human-induced IR associated with energy, weapons production, medicine, and research has changed the distribution and magnitude of these evolutionary pressures. To date, no study has systematically examined the effects of environmentally relevant doses of radiation exposure across an organismal proteome. This void in knowledge has been due, in part, to technological deficiencies that have hampered quantifiable environmentally relevant IR doses and sensitive detection of proteomic responses. Here, we describe a protocol that addresses both needs, combining quantifiable IR delivery with a reliable method to yield proteomic comparisons of control and irradiated Medaka fish. Exposures were conducted at the Savannah River Ecology Laboratory (SREL, in Aiken, SC), where fish were subsequently dissected into three tissue sets (carcasses, organs and intestines) and frozen until analysis. Tissue proteins were extracted, resolved by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE), and each sample lane was divided into ten equal portions. Following in-gel tryptic digestion, peptides released from each gel portion were identified and quantified by Liquid Chromatography-Mass Spectrometry (LC-MS/MS) to obtain the most complete, comparative study to date of proteomic responses to environmentally relevant doses of IR. This method provides a simple approach for use in ongoing epidemiologic studies of chronic exposure to environmentally relevant levels of IR and should also serve well in physiological, developmental, and toxicological studies.
Highlights
IntroductionIonizing radiation (IR), from other than natural sources, has become an aspect of daily life over the course of the last century
Ionizing radiation (IR), from other than natural sources, has become an aspect of daily life over the course of the last century. While sites such as Fukushima and Chernobyl are well-known and well documented sources of exposure to radiation, there remain over 1000 locations within the United States alone that are contaminated with radiation and have yet to be sufficiently studied to fully understand the risk to human health and to the environment
The methods described for this study provide a simple approach to detect proteomic responses to irradiation across different tissues in Medaka
Summary
Ionizing radiation (IR), from other than natural sources, has become an aspect of daily life over the course of the last century. Testing and manufacturing related to nuclear proliferation (for both energy and weapons) and rapid increases in the use of nuclear medicine [1], are becoming increasingly identified as sources of radionuclide contamination Such contamination can have long lasting effects on public health and the environment, in aquatic systems. The effects of radionuclides on organisms can vary depending on the dose and exposure time and may result in changes in morphology and functional activity, both at the cellular and system levels It is well documented, especially at high doses, that IR has detrimental effects on aquatic organisms. A recent study presented potential transgenerational bystander effects in fish and amphibian cells [11]
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