Quantitative Proteomics Reveals the Induction of Mitophagy in Tumor Necrosis Factor-α-activated (TNFα) Macrophages

Abstract

Macrophages play an important role in innate and adaptive immunity as professional phagocytes capable of internalizing and degrading pathogens to derive antigens for presentation to T cells. They also produce pro-inflammatory cytokines such as tumor necrosis factor alpha (TNF-␣) that mediate local and systemic responses and direct the development of adaptive immunity. The present work describes the use of label-free quantitative proteomics to profile the dynamic changes of proteins from resting and TNF-␣-activated mouse macrophages. These analyses revealed that TNF-␣ activation of macrophages led to the down-regulation of mitochondrial proteins and the differential regulation of several proteins involved in vesicle trafficking and immune response. Importantly, we found that the down-regulation of mitochondria proteins occurred through mitophagy and was specific to TNF-␣, as other cytokines such as IL-1␤ and IFN-␥ had no effect on mitochondria degradation. Furthermore, using a novel antigen presentation system, we observed that the induction of mitophagy by TNF-␣ enabled the processing and presentation of mitochondrial antigens at the cell surface by MHC class I molecules. These findings highlight an unsuspected role of TNF-␣ in mitophagy and expanded our understanding of the mechanisms responsible for MHC presentation of self-antigens. Molecular & Cellular

Parkinson’s Disease-Related Proteins PINK1 and Parkin Repress Mitochondrial Antigen Presentation

“…The RAW 264.7 macrophage cell line (ATCC), the H-2K b RAW cell line (Bell et al, 2013), and COS7 cells (ATCC) were cultured in DMEM (10% [v/v] fetal calf serum [FCS], penicillin [100 units/ml], and streptomycin [100 mg/ml]). The b-galactosidase-inducible HSV gB/K b -restricted HSV-2.3.2E2 CD8+ T cell hybridoma (2E2) was kindly provided by F. Carbone (University of Melbourne).…”

Presenting mitochondrial antigens: PINK1, Parkin and MDVs steal the show

“…In order to be presented on MHC class I molecules, antigens must be processed to peptides, classically by the proteasome. However, it has recently been shown that antigens can be processed via the lysosome, following autophagy or mitophagy [9,10]. Matheoud et al now show that, in addition, mitochondrial proteins can be transported to the lysosome by MDVs for subsequent presentation on MHC class I molecules.…”

High‐resolution quantitative proteome analysis reveals substantial differences between phagosomes of RAW 264.7 and bone marrow derived macrophages

“…In recent years, the cell line has also been used extensively in proteomics experiments including a large‐scale proteome 10, phagosome proteomics 4, 8, 11, responses to cytokines 12, 13, and identification of DNA receptors 14.…”