Abstract
Tumour progression locus 2 (TPL‐2) kinase mediates Toll‐like receptor (TLR) activation of ERK1/2 and p38α MAP kinases in myeloid cells to modulate expression of key cytokines in innate immunity. This study identified a novel MAP kinase‐independent regulatory function for TPL‐2 in phagosome maturation, an essential process for killing of phagocytosed microbes. TPL‐2 catalytic activity was demonstrated to induce phagosome acidification and proteolysis in primary mouse and human macrophages following uptake of latex beads. Quantitative proteomics revealed that blocking TPL‐2 catalytic activity significantly altered the protein composition of phagosomes, particularly reducing the abundance of V‐ATPase proton pump subunits. Furthermore, TPL‐2 stimulated the phosphorylation of DMXL1, a regulator of V‐ATPases, to induce V‐ATPase assembly and phagosome acidification. Consistent with these results, TPL‐2 catalytic activity was required for phagosome acidification and the efficient killing of Staphylococcus aureus and Citrobacter rodentium following phagocytic uptake by macrophages. TPL‐2 therefore controls innate immune responses of macrophages to bacteria via V‐ATPase induction of phagosome maturation.
Highlights
A key component of the innate immune response involves the killing of phagocytosed microbes, such as bacteria, by macrophages (Pauwels et al, 2017)
Real-time measurements showed that phagosome proteolysis was significantly reduced in Tpl2D270A/D270A bone marrow-derived macrophages (BMDMs) compared to WT controls (Fig 1B)
Thirty mins after bead uptake, phagosome cathepsin activity was significantly reduced by Tpl2D270A mutation (Fig 1C). These results showed that Tumour progression locus 2 (TPL-2) catalytic activity is required to increase protease activity inside phagosomes of primary macrophages
Summary
A key component of the innate immune response involves the killing of phagocytosed microbes, such as bacteria, by macrophages (Pauwels et al, 2017). The nascent phagosome is innocuous, and to kill internalised bacteria, the phagosome must mature via an ordered series of membrane fusion and fission events with endosomes and with lysosomes to form the phagolysosome, a potent microbicidal organelle (Flannagan et al, 2009). Phagosomes become increasingly acidic, highly oxidative and enriched with proteases and hydrolases that can degrade the internalised bacteria (Flannagan et al, 2009). Acidification inhibits bacterial growth (Downey et al, 1999; Ip et al, 2010), activates cathepsin proteases that degrade internalised bacteria (Yates et al, 2005) and promotes NOX2 production of reactive oxygen species (ROS) that damage bacterial proteins, lipids and nucleic acids (Savina et al, 2006)
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