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Bellutti L, Mege RM, Dang T, Chevreux G, Lignieres L, Camadro JM, Senecaut N, Terrier S, Legros V, Hamon M, Dumont J, Lelandais G
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J Proteome Res,
2023]
The simple light isotope metabolic-labeling technique relies on the in vivo biosynthesis of amino acids from U-[12C]-labeled molecules provided as the sole carbon source. The incorporation of the resulting U-[12C]-amino acids into proteins presents several key advantages for mass-spectrometry-based proteomics analysis, as it results in more intense monoisotopic ions, with a better signal-to-noise ratio in bottom-up analysis. In our initial studies, we developed the simple light isotope metabolic (SLIM)-labeling strategy using prototrophic eukaryotic microorganisms, the yeasts Candida albicans and Saccharomyces cerevisiae, as well as strains with genetic markers that lead to amino-acid auxotrophy. To extend the range of SLIM-labeling applications, we evaluated (i) the incorporation of U-[12C]-glucose into proteins of human cells grown in a complex RPMI-based medium containing the labeled molecule, considering that human cell lines require a large number of essential amino-acids to support their growth, and (ii) an indirect labeling strategy in which the nematode Caenorhabditis elegans grown on plates was fed U-[12C]-labeled bacteria (Escherichia coli) and the worm proteome analyzed for 12C incorporation into proteins. In both cases, we were able to demonstrate efficient incorporation of 12C into the newly synthesized proteins, opening the way for original approaches in quantitative proteomics.
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[
Geochim Cosmochim Acta,
1978]
The influence of diet on the distribution of carbon isotopes in animals was investigated by analyzing animals grown in the laboratory on diets of constant carbon isotopic composition. The isotopic composition of the whole body of an animal reflects the isotopic composition of its diet, but the animal is on average enriched in *13C by about 1% relative to the diet. In three of the four cases examined, the 13C enrichment of the whole body relative to the diet is balanced by a 13C depletion of the respired CO2. The isotopic relationships between the whole bodies of animals and their diets are similar for different species raised on the same diet and for the same species raised on different diets. However, the *13C values of whole bodies of individuals of a species raised on the same diet may differ by up to 2%. The relationship between the 13C/12C ratio of a tissue and the 13C/12C ratio of the diet depends both on the type of tissue and on the nature of the diet. Many of the isotopic relationships among the major biochemical fractions, namely the lipid, carbohydrate and protein fractions, are qualitatively preserved as diet carbon is incorporated into the animal. However, the difference between the *13C values of a biochemical fraction in an animal and in its diet may be as large as 3%. The *13C values of the biochemical components collagen, chitin and the insoluble organic fraction of shells, all of which are often preserved in fossil material, are related to the isotopic composition of the diet. These results indicate that it will be possible to perform dietary analysis based on the determination of the 13C/12C ratio of animal carbon. Analysis of the total animal carbon will in most cases provide a better measure of diet than the analysis of individual tissues, biochemical fractions, or biochemical components. The limits of accuracy of this method will generally restrict its application to situations in which the diet is derived from sources with relatively large differences in their *13C values, such as terrestrial vs aquatic organisms or C3 vs C4 plants. This method should be applicable to fossil
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[
PLoS One,
2016]
A wealth of studies has investigated how chemical sensitivity is affected by temperature, however, almost always under different constant rather than more realistic fluctuating regimes. Here we compared how the nematode Caenorhabditis elegans responds to copper at constant temperatures (8-24C) and under fluctuation conditions of low (+/-4C) and high (+/-8C) amplitude (averages of 12, 16, 20C and 16C respectively). The DEBkiss model was used to interpret effects on energy budgets. Increasing constant temperature from 12-24C reduced time to first egg, life-span and population growth rates consistent with temperature driven metabolic rate change. Responses at 8C did not, however, accord with this pattern (including a deviation from the Temperature Size Rule), identifying a cold stress effect. High amplitude variation and low amplitude variation around a mean temperature of 12C impacted reproduction and body size compared to nematodes kept at the matching average constant temperatures. Copper exposure affected reproduction, body size and life-span and consequently population growth. Sensitivity to copper (EC50 values), was similar at intermediate temperatures (12, 16, 20C) and higher at 24C and especially the innately stressful 8C condition. Temperature variation did not increase copper sensitivity. Indeed under variable conditions including time at the stressful 8C condition, sensitivity was reduced. DEBkiss identified increased maintenance costs and increased assimilation as possible mechanisms for cold and higher copper concentration effects. Model analysis of combined variable temperature effects, however, demonstrated no additional joint stressor response. Hence, concerns that exposure to temperature fluctuations may sensitise species to co-stressor effects seem unfounded in this case.
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[
J Radiat Res,
2017]
Radiation may affect essential functions and behaviors such as locomotion, feeding, learning and memory. Although whole-body irradiation has been shown to reduce motility in the nematode Caenorhabditis elegans, the detailed mechanism responsible for this effect remains unknown. Targeted irradiation of the nerve ring responsible for sensory integration and information processing would allow us to determine whether the reduction of motility following whole-body irradiation reflects effects on the central nervous system or on the muscle cells themselves. We therefore addressed this issue using a collimating microbeam system. However, radiation targeting requires the animal to be immobilized, and previous studies have anesthetized animals to prevent their movement, thus making it impossible to assess their locomotion immediately after irradiation. We developed a method in which the animal was enclosed in a straight, microfluidic channel in a polydimethylsiloxane chip to inhibit free motion during irradiation, thus allowing locomotion to be observed immediately after irradiation. The head region (including the central nervous system), mid region around the intestine and uterus, and tail region were targeted independently. Each region was irradiated with 12 000 carbon ions (12C; 18.3 MeV/u; linear energy transfer = 106.4 keV/m), corresponding to 500 Gy at a 20 m region. Motility was significantly decreased by whole-body irradiation, but not by irradiation of any of the individual regions, including the central nervous system. This suggests that radiation inhibits locomotion by a whole-body mechanism, potentially involving motoneurons and/or body-wall muscle cells, rather than affecting motor control via the central nervous system and the stimulation response.