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

1
Institute of Clinical Chemistry, Hannover Medical School, 30625 Hannover, Germany
2
Institute of Toxicology, Hannover Medical School, 30625 Hannover, Germany
3
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; https://doi.org/10.3390/ijms20051241
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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Abstract

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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