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DiDBiT-TMT: A Novel Method to Quantify Changes in the Proteomic Landscape Induced by Neural Plasticity

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Download Hi-Res ImageDownload to MS-PowerPointCite This:J. Proteome Res. 2024, 23, 11, 4878-4895
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    DiDBiT-TMT: A Novel Method to Quantify Changes in the Proteomic Landscape Induced by Neural Plasticity
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    • Mariam Gamaleldin
      Mariam Gamaleldin
      Danish Research Institute of Translational Neuroscience DANDRITE-Nordic EMBL Partnership for Molecular Medicine, Aarhus University, Aarhus C 8000, Denmark
      Department of Molecular Biology and Genetics, Aarhus C 8000, Denmark
      School of Biotechnology, Nile University, Giza 12588, Egypt
      More by Mariam Gamaleldin
      Orcidhttps://orcid.org/0000-0001-6281-7102
    • Nam-Kyung Yu
      Nam-Kyung Yu
      Departments of Molecular Medicine and Neurobiology, the Scripps Research Institute, La Jolla 92037, California, United States
      More by Nam-Kyung Yu
      Orcidhttps://orcid.org/0000-0002-7432-7394
    • Jolene K Diedrich
      Jolene K Diedrich
      Departments of Molecular Medicine and Neurobiology, the Scripps Research Institute, La Jolla 92037, California, United States
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      Orcidhttps://orcid.org/0000-0001-6489-4558
    • Yuanhui Ma
      Yuanhui Ma
      Departments of Molecular Medicine and Neurobiology, the Scripps Research Institute, La Jolla 92037, California, United States
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    • Anne Wienand
      Anne Wienand
      The Danish National Research Foundation Center of Excellence PROMEMO, Aarhus University, Aarhus C 8000, Denmark
      Danish Research Institute of Translational Neuroscience DANDRITE-Nordic EMBL Partnership for Molecular Medicine, Aarhus University, Aarhus C 8000, Denmark
      Department of Molecular Biology and Genetics, Aarhus C 8000, Denmark
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    • Daniel B. McClatchy
      Daniel B. McClatchy
      Departments of Molecular Medicine and Neurobiology, the Scripps Research Institute, La Jolla 92037, California, United States
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      Orcidhttps://orcid.org/0000-0002-0288-5645
    • Anders Nykjaer
      Anders Nykjaer
      The Danish National Research Foundation Center of Excellence PROMEMO, Aarhus University, Aarhus C 8000, Denmark
      Danish Research Institute of Translational Neuroscience DANDRITE-Nordic EMBL Partnership for Molecular Medicine, Aarhus University, Aarhus C 8000, Denmark
      Department of Biomedicine, Aarhus University, Aarhus C 8000, Denmark
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    • Sadegh Nabavi*
      Sadegh Nabavi
      The Danish National Research Foundation Center of Excellence PROMEMO, Aarhus University, Aarhus C 8000, Denmark
      Danish Research Institute of Translational Neuroscience DANDRITE-Nordic EMBL Partnership for Molecular Medicine, Aarhus University, Aarhus C 8000, Denmark
      Department of Molecular Biology and Genetics, Aarhus C 8000, Denmark
      *Email: [email protected]
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    • John R. Yates III*
      John R. Yates, III
      Departments of Molecular Medicine and Neurobiology, the Scripps Research Institute, La Jolla 92037, California, United States
      *Email: [email protected]
      More by John R. Yates, III
      Orcidhttps://orcid.org/0000-0001-5267-1672
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    Journal of Proteome Research

    Cite this: J. Proteome Res. 2024, 23, 11, 4878–4895
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    https://doi.org/10.1021/acs.jproteome.4c00180
    Published October 7, 2024
    Copyright © 2024 American Chemical Society
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    Abstract

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    Direct detection of biotinylated proteins (DiDBiT) is a proteomic method that can enrich and detect newly synthesized proteins (NSPs) labeled with bio-orthogonal amino acids with 20-fold improved detectability compared to conventional methods. However, DiDBiT has currently been used to compare only two conditions per experiment. Here, we present DiDBiT-TMT, a method that can be used to quantify NSPs across many conditions and replicates in the same experiment by combining isobaric tandem mass tagging (TMT) with DiDBiT. We applied DiDBiT-TMT to brain slices to determine changes in the de novo proteome that occur after inducing chemical long-term potentiation (cLTP) or treatment with the neuromodulator norepinephrine. We successfully demonstrated DiDBiT-TMT’s capacity to quantitatively compare up to 9 samples in parallel. We showed that there is a minimal overlap among NSPs that are differentially expressed in cLTP-treated organotypic brain slices, norepinephrine-treated organotypic brain slices, and organotypic slices undergoing combinatorial treatment with norepinephrine and cLTP. Our results point to the possible divergence of the molecular mechanisms underlying these treatments and showcase the applicability of DiDBiT-TMT for studying neurobiology.

    Copyright © 2024 American Chemical Society

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    Supporting Information

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    The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acs.jproteome.4c00180.

    • (Figure S1) pSER845 quantification across experimental conditions and (Figure S2) raw images of the Western blots (PDF)

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    Journal of Proteome Research

    Cite this: J. Proteome Res. 2024, 23, 11, 4878–4895
    Click to copy citationCitation copied!
    https://doi.org/10.1021/acs.jproteome.4c00180
    Published October 7, 2024
    Copyright © 2024 American Chemical Society
    Request reuse permissions

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