DIA-based systems biology approach unveils E3 ubiquitin ligase-dependent responses to a metabolic shift
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DIA-based systems biology approach unveils E3 ubiquitin ligase-dependent responses to a metabolic shift. / Karayel, Ozge; Michaelis, André C; Mann, Matthias; Schulman, Brenda A; Langlois, Christine R.
In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 117, No. 51, 22.12.2020, p. 32806-32815.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - DIA-based systems biology approach unveils E3 ubiquitin ligase-dependent responses to a metabolic shift
AU - Karayel, Ozge
AU - Michaelis, André C
AU - Mann, Matthias
AU - Schulman, Brenda A
AU - Langlois, Christine R
N1 - Copyright © 2020 the Author(s). Published by PNAS.
PY - 2020/12/22
Y1 - 2020/12/22
N2 - The yeast Saccharomyces cerevisiae is a powerful model system for systems-wide biology screens and large-scale proteomics methods. Nearly complete proteomics coverage has been achieved owing to advances in mass spectrometry. However, it remains challenging to scale this technology for rapid and high-throughput analysis of the yeast proteome to investigate biological pathways on a global scale. Here we describe a systems biology workflow employing plate-based sample preparation and rapid, single-run, data-independent mass spectrometry analysis (DIA). Our approach is straightforward, easy to implement, and enables quantitative profiling and comparisons of hundreds of nearly complete yeast proteomes in only a few days. We evaluate its capability by characterizing changes in the yeast proteome in response to environmental perturbations, identifying distinct responses to each of them and providing a comprehensive resource of these responses. Apart from rapidly recapitulating previously observed responses, we characterized carbon source-dependent regulation of the GID E3 ligase, an important regulator of cellular metabolism during the switch between gluconeogenic and glycolytic growth conditions. This unveiled regulatory targets of the GID ligase during a metabolic switch. Our comprehensive yeast system readout pinpointed effects of a single deletion or point mutation in the GID complex on the global proteome, allowing the identification and validation of targets of the GID E3 ligase. Moreover, this approach allowed the identification of targets from multiple cellular pathways that display distinct patterns of regulation. Although developed in yeast, rapid whole-proteome-based readouts can serve as comprehensive systems-level assays in all cellular systems.
AB - The yeast Saccharomyces cerevisiae is a powerful model system for systems-wide biology screens and large-scale proteomics methods. Nearly complete proteomics coverage has been achieved owing to advances in mass spectrometry. However, it remains challenging to scale this technology for rapid and high-throughput analysis of the yeast proteome to investigate biological pathways on a global scale. Here we describe a systems biology workflow employing plate-based sample preparation and rapid, single-run, data-independent mass spectrometry analysis (DIA). Our approach is straightforward, easy to implement, and enables quantitative profiling and comparisons of hundreds of nearly complete yeast proteomes in only a few days. We evaluate its capability by characterizing changes in the yeast proteome in response to environmental perturbations, identifying distinct responses to each of them and providing a comprehensive resource of these responses. Apart from rapidly recapitulating previously observed responses, we characterized carbon source-dependent regulation of the GID E3 ligase, an important regulator of cellular metabolism during the switch between gluconeogenic and glycolytic growth conditions. This unveiled regulatory targets of the GID ligase during a metabolic switch. Our comprehensive yeast system readout pinpointed effects of a single deletion or point mutation in the GID complex on the global proteome, allowing the identification and validation of targets of the GID E3 ligase. Moreover, this approach allowed the identification of targets from multiple cellular pathways that display distinct patterns of regulation. Although developed in yeast, rapid whole-proteome-based readouts can serve as comprehensive systems-level assays in all cellular systems.
KW - Carbon/metabolism
KW - Culture Media
KW - Fructose-Bisphosphatase/metabolism
KW - Glucose/metabolism
KW - Malate Dehydrogenase/metabolism
KW - Mass Spectrometry/methods
KW - Point Mutation
KW - Pyruvate Decarboxylase/metabolism
KW - Saccharomyces cerevisiae/growth & development
KW - Saccharomyces cerevisiae Proteins/genetics
KW - Sodium-Potassium-Exchanging ATPase/metabolism
KW - Stress, Physiological
KW - Systems Biology/methods
KW - Ubiquitin-Protein Ligases/genetics
KW - Workflow
U2 - 10.1073/pnas.2020197117
DO - 10.1073/pnas.2020197117
M3 - Journal article
C2 - 33288721
VL - 117
SP - 32806
EP - 32815
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
SN - 0027-8424
IS - 51
ER -
ID: 259832837