Global protein phosphorylation dynamics during deoxynivalenol-induced ribotoxic stress response in the macrophage

Xiao Pan, Douglas A. Whitten, Ming Wu, Christina Chan, Curtis G. Wilkerson, James J. Pestka

Research output: Contribution to journalArticle

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Abstract

Deoxynivalenol (DON), a trichothecene mycotoxin produced by Fusarium that commonly contaminates food, is capable of activating mononuclear phagocytes of the innate immune system via a process termed the ribotoxic stress response (RSR). To encapture global signaling events mediating RSR, we quantified the early temporal (≤ 30. min) phosphoproteome changes that occurred in RAW 264.7 murine macrophage during exposure to a toxicologically relevant concentration of DON (250. ng/mL). Large-scale phosphoproteomic analysis employing stable isotope labeling of amino acids in cell culture (SILAC) in conjunction with titanium dioxide chromatography revealed that DON significantly upregulated or downregulated phosphorylation of 188 proteins at both known and yet-to-be functionally characterized phosphosites. DON-induced RSR is extremely complex and goes far beyond its prior known capacity to inhibit translation and activate MAPKs. Transcriptional regulation was the main target during early DON-induced RSR, covering over 20% of the altered phosphoproteins as indicated by Gene Ontology annotation and including transcription factors/cofactors and epigenetic modulators. Other biological processes impacted included cell cycle, RNA processing, translation, ribosome biogenesis, monocyte differentiation and cytoskeleton organization. Some of these processes could be mediated by signaling networks involving MAPK-, NFκB-, AKT- and AMPK-linked pathways. Fuzzy c-means clustering revealed that DON-regulated phosphosites could be discretely classified with regard to the kinetics of phosphorylation/dephosphorylation. The cellular response networks identified provide a template for further exploration of the mechanisms of trichothecenemycotoxins and other ribotoxins, and ultimately, could contribute to improved mechanism-based human health risk assessment.

LanguageEnglish (US)
Pages201-211
Number of pages11
JournalToxicology and Applied Pharmacology
Volume268
Issue number2
DOIs
StatePublished - Apr 5 2013

Profile

Phosphorylation
Macrophages
Proteins
Trichothecenes
Isotope Labeling
Molecular Sequence Annotation
Biological Phenomena
Gene Ontology
AMP-Activated Protein Kinases
Mycotoxins
Immune system
Health risks
Phosphoproteins
Fusarium
Phagocytes
Chromatography
Cytoskeleton
Ribosomes
Cell culture
Epigenomics

Keywords

  • Deoxynivalenol
  • Phosphorylation
  • Quantitative proteomics
  • Ribotoxic stress response
  • Trichothecene mycotoxin

ASJC Scopus subject areas

  • Pharmacology
  • Toxicology

Cite this

Global protein phosphorylation dynamics during deoxynivalenol-induced ribotoxic stress response in the macrophage. / Pan, Xiao; Whitten, Douglas A.; Wu, Ming; Chan, Christina; Wilkerson, Curtis G.; Pestka, James J.

In: Toxicology and Applied Pharmacology, Vol. 268, No. 2, 05.04.2013, p. 201-211.

Research output: Contribution to journalArticle

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abstract = "Deoxynivalenol (DON), a trichothecene mycotoxin produced by Fusarium that commonly contaminates food, is capable of activating mononuclear phagocytes of the innate immune system via a process termed the ribotoxic stress response (RSR). To encapture global signaling events mediating RSR, we quantified the early temporal (≤ 30. min) phosphoproteome changes that occurred in RAW 264.7 murine macrophage during exposure to a toxicologically relevant concentration of DON (250. ng/mL). Large-scale phosphoproteomic analysis employing stable isotope labeling of amino acids in cell culture (SILAC) in conjunction with titanium dioxide chromatography revealed that DON significantly upregulated or downregulated phosphorylation of 188 proteins at both known and yet-to-be functionally characterized phosphosites. DON-induced RSR is extremely complex and goes far beyond its prior known capacity to inhibit translation and activate MAPKs. Transcriptional regulation was the main target during early DON-induced RSR, covering over 20{\%} of the altered phosphoproteins as indicated by Gene Ontology annotation and including transcription factors/cofactors and epigenetic modulators. Other biological processes impacted included cell cycle, RNA processing, translation, ribosome biogenesis, monocyte differentiation and cytoskeleton organization. Some of these processes could be mediated by signaling networks involving MAPK-, NFκB-, AKT- and AMPK-linked pathways. Fuzzy c-means clustering revealed that DON-regulated phosphosites could be discretely classified with regard to the kinetics of phosphorylation/dephosphorylation. The cellular response networks identified provide a template for further exploration of the mechanisms of trichothecenemycotoxins and other ribotoxins, and ultimately, could contribute to improved mechanism-based human health risk assessment.",
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