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Measuring Food Intake and Nutrient Absorption in Caenorhabditis elegans.

Gomez-Amaro RL, Valentine ER, Carretero M, LeBoeuf SE, Rangaraju S, Broaddus CD, Solis GM, Williamson JR, Petrascheck M - Genetics (2015)

Bottom Line: Caenorhabditis elegans has emerged as a powerful model to study the genetics of feeding, food-related behaviors, and metabolism.We show that serotonin-increased feeding leads to increased protein synthesis in a SER-7-dependent manner, including proteins known to promote aging.Protein content in the food has recently emerged as critical factor in determining how food composition affects aging and health.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical Physiology, The Scripps Research Institute, La Jolla, California 92037 Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, California 92037 Department of Molecular and Cellular Neuroscience, The Scripps Research Institute, La Jolla, California 92037.

No MeSH data available.


Pulse-feeding assay. (A) Whole worm lysate analyzed on the mass spectrometer, with sample data shown in Figure 4, B and C. Each peptide in the spectra has a “light,” “medium,” and “heavy” component. The intensity of the light component depends on the amount of each peptide present before the pulse-labeling period. The intensity of the medium component depends on the amount of ingested food after the start of the pulse-labeling period. The heavy component serves as a standard sample. (B) Correlation plot of fraction-labeled values for wild-type N2 (x-axis) vs.tph-1 mutant worms (y-axis) from L4:D1. (C) Correlation plot of fraction-labeled values by protein for wild-type N2 treated with water (x-axis) or serotonin (y-axis, 5 mM) from D5:D7. (D) Correlation plot of fraction-labeled values for ser-7 mutant worms grown with water (x-axis) or treated with serotonin (y-axis, 5 mM) from D5:D7. (E) Correlation plot of fraction-labeled values for wild-type N2 (y-axis) and ser-7 mutants (x-axis) treated with water from D5:D7. (F) Histogram of protein level values for wild-type N2 and ser-7 mutants treated with water or serotonin (5 mM).
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fig5: Pulse-feeding assay. (A) Whole worm lysate analyzed on the mass spectrometer, with sample data shown in Figure 4, B and C. Each peptide in the spectra has a “light,” “medium,” and “heavy” component. The intensity of the light component depends on the amount of each peptide present before the pulse-labeling period. The intensity of the medium component depends on the amount of ingested food after the start of the pulse-labeling period. The heavy component serves as a standard sample. (B) Correlation plot of fraction-labeled values for wild-type N2 (x-axis) vs.tph-1 mutant worms (y-axis) from L4:D1. (C) Correlation plot of fraction-labeled values by protein for wild-type N2 treated with water (x-axis) or serotonin (y-axis, 5 mM) from D5:D7. (D) Correlation plot of fraction-labeled values for ser-7 mutant worms grown with water (x-axis) or treated with serotonin (y-axis, 5 mM) from D5:D7. (E) Correlation plot of fraction-labeled values for wild-type N2 (y-axis) and ser-7 mutants (x-axis) treated with water from D5:D7. (F) Histogram of protein level values for wild-type N2 and ser-7 mutants treated with water or serotonin (5 mM).

Mentions: To understand how we identify individual peptides by mass spectrometry, it is necessary to address the complexity of the peptide samples. For each peptide, three isotope enrichment patterns are possible: 100% 14N-enriched (L), if it was synthesized before the pulse; 50% 15N-enriched (M), if it was synthesized during the pulse; or 100% 15N-enriched (H), if it was part of the external standard (Figure 4, A–C). Thus, each peptide in the sample can have three components in its isotope distribution in the mass spectra. Database search engines cannot identify peptides from 50% 15N-enriched parent ions and, as a consequence, only the identification of the 100% 14N- and 100% 15N-labeled peptides was carried out using MASCOT with searches for both E. coli and C. elegans (Eng et al. 1994). Because the light, medium, and heavy peptides coelute during liquid chromatography, the MS1 scan for the m/z range spanning all three labeled species can be extracted (L, M, and H) (Figure 5A). Misidentifications are readily identified because the spacing of the 14N and 15N peaks does not match the number of nitrogen atoms in the sequence. Thus, we can ensure correct identification of each peptide, despite the complexity of the isotope distribution with three components.


Measuring Food Intake and Nutrient Absorption in Caenorhabditis elegans.

Gomez-Amaro RL, Valentine ER, Carretero M, LeBoeuf SE, Rangaraju S, Broaddus CD, Solis GM, Williamson JR, Petrascheck M - Genetics (2015)

Pulse-feeding assay. (A) Whole worm lysate analyzed on the mass spectrometer, with sample data shown in Figure 4, B and C. Each peptide in the spectra has a “light,” “medium,” and “heavy” component. The intensity of the light component depends on the amount of each peptide present before the pulse-labeling period. The intensity of the medium component depends on the amount of ingested food after the start of the pulse-labeling period. The heavy component serves as a standard sample. (B) Correlation plot of fraction-labeled values for wild-type N2 (x-axis) vs.tph-1 mutant worms (y-axis) from L4:D1. (C) Correlation plot of fraction-labeled values by protein for wild-type N2 treated with water (x-axis) or serotonin (y-axis, 5 mM) from D5:D7. (D) Correlation plot of fraction-labeled values for ser-7 mutant worms grown with water (x-axis) or treated with serotonin (y-axis, 5 mM) from D5:D7. (E) Correlation plot of fraction-labeled values for wild-type N2 (y-axis) and ser-7 mutants (x-axis) treated with water from D5:D7. (F) Histogram of protein level values for wild-type N2 and ser-7 mutants treated with water or serotonin (5 mM).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig5: Pulse-feeding assay. (A) Whole worm lysate analyzed on the mass spectrometer, with sample data shown in Figure 4, B and C. Each peptide in the spectra has a “light,” “medium,” and “heavy” component. The intensity of the light component depends on the amount of each peptide present before the pulse-labeling period. The intensity of the medium component depends on the amount of ingested food after the start of the pulse-labeling period. The heavy component serves as a standard sample. (B) Correlation plot of fraction-labeled values for wild-type N2 (x-axis) vs.tph-1 mutant worms (y-axis) from L4:D1. (C) Correlation plot of fraction-labeled values by protein for wild-type N2 treated with water (x-axis) or serotonin (y-axis, 5 mM) from D5:D7. (D) Correlation plot of fraction-labeled values for ser-7 mutant worms grown with water (x-axis) or treated with serotonin (y-axis, 5 mM) from D5:D7. (E) Correlation plot of fraction-labeled values for wild-type N2 (y-axis) and ser-7 mutants (x-axis) treated with water from D5:D7. (F) Histogram of protein level values for wild-type N2 and ser-7 mutants treated with water or serotonin (5 mM).
Mentions: To understand how we identify individual peptides by mass spectrometry, it is necessary to address the complexity of the peptide samples. For each peptide, three isotope enrichment patterns are possible: 100% 14N-enriched (L), if it was synthesized before the pulse; 50% 15N-enriched (M), if it was synthesized during the pulse; or 100% 15N-enriched (H), if it was part of the external standard (Figure 4, A–C). Thus, each peptide in the sample can have three components in its isotope distribution in the mass spectra. Database search engines cannot identify peptides from 50% 15N-enriched parent ions and, as a consequence, only the identification of the 100% 14N- and 100% 15N-labeled peptides was carried out using MASCOT with searches for both E. coli and C. elegans (Eng et al. 1994). Because the light, medium, and heavy peptides coelute during liquid chromatography, the MS1 scan for the m/z range spanning all three labeled species can be extracted (L, M, and H) (Figure 5A). Misidentifications are readily identified because the spacing of the 14N and 15N peaks does not match the number of nitrogen atoms in the sequence. Thus, we can ensure correct identification of each peptide, despite the complexity of the isotope distribution with three components.

Bottom Line: Caenorhabditis elegans has emerged as a powerful model to study the genetics of feeding, food-related behaviors, and metabolism.We show that serotonin-increased feeding leads to increased protein synthesis in a SER-7-dependent manner, including proteins known to promote aging.Protein content in the food has recently emerged as critical factor in determining how food composition affects aging and health.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical Physiology, The Scripps Research Institute, La Jolla, California 92037 Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, California 92037 Department of Molecular and Cellular Neuroscience, The Scripps Research Institute, La Jolla, California 92037.

No MeSH data available.