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Proteome and Phosphoproteome Analysis in TNF Long Term-Exposed Primary Human Monocytes

Institute of Clinical Chemistry, Hannover Medical School, 30625 Hannover, Germany
Institute of Toxicology, Hannover Medical School, 30625 Hannover, Germany
Core Unit Proteomics, Hannover Medical School, 30625 Hannover, Germany
Author to whom correspondence should be addressed.
These authors contributed equally to this work.
These authors contributed equally to this work.
Int. J. Mol. Sci. 2019, 20(5), 1241;
Received: 11 February 2019 / Revised: 27 February 2019 / Accepted: 6 March 2019 / Published: 12 March 2019
(This article belongs to the Special Issue Tumor Necrosis Factor (TNF))
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To better understand the inflammation-associated mechanisms modulating and terminating tumor necrosis factor (TNF-)induced signal transduction and the development of TNF tolerance, we analyzed both the proteome and the phosphoproteome in TNF long term-incubated (i.e., 48 h) primary human monocytes using liquid chromatography-mass spectrometry. Our analyses revealed the presence of a defined set of proteins characterized by reproducible changes in expression and phosphorylation patterns in long term TNF-treated samples. In total, 148 proteins and 569 phosphopeptides were significantly regulated (103 proteins increased, 45 proteins decreased; 377 peptides with increased and 192 peptides with decreased phosphorylation). A variety of these proteins are associated with the non-canonical nuclear factor κB (NF-κB) pathway (nuclear factor κB (NFKB) 2, v-rel reticuloendotheliosis viral oncogene homolog (REL) B, indolamin-2,3-dioxygenase (IDO), kynureninase (KYNU)) or involved in the negative regulation of the canonical NF-κB system. Within the phosphopeptides, binding motifs for specific kinases were identified. Glycogen synthase kinase (GSK) 3 proved to be a promising candidate, since it targets NF-κB inhibiting factors, such as CCAAT/enhancer binding protein (C/EBP) β. Our experiments demonstrate that both proteome and phosphoproteome analysis can be effectively applied to study protein/phosphorylation patterns of primary monocytes. These results provide new regulatory candidates and evidence for a complex network of specific but synergistically acting/cooperating mechanisms enabling the affected cells to resist sustained TNF exposure and resulting in the resolution of inflammation. View Full-Text
Keywords: proteomics; phosphoproteomics; TNF long term exposure; monocytes; NF-κB proteomics; phosphoproteomics; TNF long term exposure; monocytes; NF-κB

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Welz, B.; Bikker, R.; Junemann, J.; Christmann, M.; Neumann, K.; Weber, M.; Hoffmeister, L.; Preuß, K.; Pich, A.; Huber, R.; Brand, K. Proteome and Phosphoproteome Analysis in TNF Long Term-Exposed Primary Human Monocytes. Int. J. Mol. Sci. 2019, 20, 1241.

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