2025-06-02 パシフィック・ノースウェスト国立研究所(PNNL)
<関連情報>
- https://www.pnnl.gov/publications/researchers-develop-new-integrative-and-automated-approach-identify-multiple-types
- https://www.mcponline.org/article/S1535-9476(24)00171-3/fulltext
統合的マルチPTMプロテオミクスによるサイトカイン処理膵β細胞における動的なグローバル、レドックス、リン酸化、アセチル化制御の解明 Integrative Multi-PTM Proteomics Reveals Dynamic Global, Redox, Phosphorylation, and Acetylation Regulation in Cytokine-Treated Pancreatic Beta Cells
Austin Gluth ∙ Xiaolu Li ∙ Marina A. Gritsenko ∙ … ∙ Bin Yang ∙ Wei-Jun Qian ∙ Tong Zhang
Molecular & Cellular Proteomics Published:November 14, 2024
DOI:https://doi.org/10.1016/j.mcpro.2024.100881
Graphical Abstract
Highlights
- SP3 is compatible with proteomics analysis of cysteine thiol oxidation.
- An automated SP3-based workflow facilitates high-throughput multi-PTM profiling.
- Cytokines induces time-dependent regulation of protein abundance and PTMs.
- Interplay of thiol oxidation, phosphorylation, and acetylation on selected proteins.
Abstract
Studying regulation of protein function at a systems level necessitates an understanding of the interplay among diverse posttranslational modifications (PTMs). A variety of proteomics sample processing workflows are currently used to study specific PTMs but rarely characterize multiple types of PTMs from the same sample inputs. Method incompatibilities and laborious sample preparation steps complicate large-scale physiological investigations and can lead to variations in results. The single-pot, solid-phase–enhanced sample preparation (SP3) method for sample cleanup is compatible with different lysis buffers and amenable to automation, making it attractive for high-throughput multi-PTM profiling. Herein, we describe an integrative SP3 workflow for multiplexed quantification of protein abundance, cysteine thiol oxidation, phosphorylation, and acetylation. The broad applicability of this approach is demonstrated using cell and tissue samples, and its utility for studying interacting regulatory networks is highlighted in a time-course experiment of cytokine-treated β-cells. We observed a swift response in the global regulation of protein abundances consistent with rapid activation of JAK-STAT and NF-κB signaling pathways. Regulators of these pathways as well as proteins involved in their target processes displayed multi-PTM dynamics indicative of complex cellular response stages: acute, adaptation, and chronic (prolonged stress). PARP14, a negative regulator of JAK-STAT, had multiple colocalized PTMs that may be involved in intraprotein regulatory crosstalk. Our workflow provides a high-throughput platform that can profile multi-PTMomes from the same sample set, which is valuable in unraveling the functional roles of PTMs and their co-regulation.
