Modulation of the cell membrane lipid milieu by peroxisomal ß-oxidation induces Rho1 signaling to trigger inflammatory responses.

Phagocytosis, signal transduction, and inflammatory responses require changes in lipid metabolism. Peroxisomes have key roles in fatty acid homeostasis and in regulating immune function. We find that Drosophila macrophages lacking peroxisomes have perturbed lipid profiles, which reduce host survival after infection. Using lipidomic, transcriptomic, and genetic screens, we determine that peroxisomes contribute to the cell membrane glycerophospholipid composition necessary to induce Rho1-dependent signals, which drive cytoskeletal remodeling during macrophage activation. Loss of peroxisome function increases membrane phosphatidic acid (PA) and recruits RhoGAPp190 during infection, inhibiting Rho1-mediated responses. Peroxisome-glycerophospholipid-Rho1 signaling also controls cytoskeleton remodeling in mouse immune cells. While high levels of PA in cells without peroxisomes inhibit inflammatory phenotypes, large numbers of peroxisomes and low amounts of cell membrane PA are features of immune cells from patients with inflammatory Kawasaki disease and juvenile idiopathic arthritis. Our findings reveal potential metabolic markers and therapeutic targets for immune diseases and metabolic disorders.

Knockdown of PEX16 Induces Autophagic Degradation of Peroxisomes.

Peroxisome abundance is regulated by homeostasis between the peroxisomal biogenesis and degradation processes. Peroxin 16 (PEX16) is a peroxisomal protein involved in trafficking membrane proteins for de novo peroxisome biogenesis. The present study demonstrates that PEX16 also modulates peroxisome abundance through pexophagic degradation. PEX16 knockdown in human retinal pigment epithelial-1 cells decreased peroxisome abundance and function, represented by reductions in the expression of peroxisome membrane protein ABCD3 and the levels of cholesterol and plasmalogens, respectively. The activation of pexophagy under PEX16 knockdown was shown by (i) abrogated peroxisome loss under PEX16 knockdown in autophagy-deficient ATG5 knockout cell lines, and (ii) increased autophagy flux and co-localization of p62-an autophagy adaptor protein-with ABCD3 in the presence of the autophagy inhibitor chloroquine. However, the levels of cholesterol and plasmalogens did not recover despite the restoration of peroxisome abundance following chloroquine treatment. Thus, PEX16 is indispensable for maintaining peroxisome homeostasis by regulating not only the commonly known biogenesis pathway but also the autophagic degradation of peroxisomes.

Pharmacological inhibition of catalase induces peroxisome leakage and suppression of LPS induced inflammatory response in Raw 264.7 cell.

Peroxisomes are metabolically active organelles which are known to exert anti-inflammatory effects especially associated with the synthesis of mediators of inflammation resolution. However, the role of catalase and effects of peroxisome derived reactive oxygen species (ROS) caused by lipid peroxidation through 4-hydroxy-2-nonenal (4-HNE) on lipopolysaccharide (LPS) mediated inflammatory pathway are largely unknown. Here, we show that inhibition of catalase by 3-aminotriazole (3-AT) results in the generation of peroxisomal ROS, which contribute to leaky peroxisomes in RAW264.7 cells. Leaky peroxisomes cause the release of matrix proteins to the cytosol, which are degraded by ubiquitin proteasome system. Furthermore, 3-AT promotes the formation of 4HNE-IκBα adduct which directly interferes with LPS induced NF-κB activation. Even though, a selective degradation of peroxisome matrix proteins and formation of 4HNE- IκBα adduct are not directly related with each other, both of them are could be the consequences of lipid peroxidation occurring at the peroxisome membrane.

Dimethyloxaloylglycine induces pexophagy in a HIF-2α dependent manner involving autophagy receptor p62.

Peroxisomes are metabolically active oxygen demanding organelles with a high abundance of oxidases making it vulnerable to low oxygen levels such as hypoxic conditions. However, the exact mechanism of peroxisome degradation in hypoxic condition remains elusive. In order to study the mechanism of peroxisome degradation in hypoxic condition, we use Dimethyloxaloylglycine (DMOG), a cell-permeable prolyl-4-hydroxylase inhibitor, which mimics hypoxic condition by stabilizing hypoxia-inducible factors. Here we report that DMOG degraded peroxisomes by selectively activating pexophagy in a HIF-2α dependent manner involving autophagy receptor p62. Furthermore, DMOG not only increased peroxisome turnover by pexophagy but also reduced HIF-2α dependent peroxisome proliferation at the transcriptional level. Taken together, our data suggest that hypoxic condition is a negative regulator for peroxisome abundance through increasing pexophagy and decreasing peroxisome proliferation in HIF-2α dependent manner.