JNK and Ceramide Kinase Govern the Biogenesis of Lipid Droplets through Activation of Group IVA Phospholipase A2

Abstract

The biogenesis of lipid droplets (LD) induced by serum depends on group IVA phospholipase A 2 (cPLA 2 ␣). This work dissects the pathway leading to cPLA 2 ␣ activation and LD biogenesis. Both processes were Ca 2؉ -independent, as they took place after pharmacological blockade of Ca 2؉ transients elicited by serum or chelation with 1,2-bis(2-aminophenoxy)ethane-N,N,N,N-tetraacetic acid tetrakis(acetoxymethyl ester). The single mutation D43N in cPLA 2 ␣, which abrogates its Ca 2؉ binding capacity and translocation to membranes, did not affect enzyme activation and formation of LD. In contrast, the mutation S505A did not affect membrane relocation of the enzyme in response to Ca 2؉ but prevented its phosphorylation, activation, and the appearance of LD. Expression of specific activators of different mitogen-activated protein kinases showed that phosphorylation of cPLA 2 ␣ at Ser-505 is due to JNK. This was confirmed by pharmacological inhibition and expression of a dominant-negative form of the upstream activator MEKK1. LD biogenesis was accompanied by increased synthesis of ceramide 1-phosphate. Overexpression of its synthesizing enzyme ceramide kinase increased phosphorylation of cPLA 2 ␣ at Ser-505 and formation of LD, and its down-regulation blocked the phosphorylation of cPLA 2 ␣ and LD biogenesis. These results demonstrate that LD biogenesis induced by serum is regulated by JNK and ceramide kinase.

Subcellular Localization and Role of Lipin-1 in Human Macrophages

“…In this regard, we have recently described the JNK-dependent phosphorylation activation of cPLA 2 a in human monocytederived macrophages (39). Importantly, in other series of experiments we also found that JNK activation is required for LD production in serum-activated cells (36). Whether JNK activation provides a molecular link between cPLA 2 a activation and proper LD formation under lipin-1 modulation is currently under active investigation in our laboratory.…”

“…We have recently shown that cPLA 2 a regulates the biogenesis of LDs in a model cell system (34)(35)(36). Interestingly, previous work has demonstrated that, under certain conditions, cPLA 2 a activation may occur downstream of a Mg 2+ -dependent PAP enzyme via DAG generation (16,18).…”

“…Leukocyte LDs have been shown to contain esterified AA and to associate to proteins that are implicated in the generation of eicosanoids (29)(30)(31)(32)(33). In turn, we have recently described that cytosolic phospholipase A 2 a (also known as group IVA phospholipase A 2 , cPLA 2 a), that is, the major effector of AA release that leads to PG and leukotriene production, is involved in the biogenesis of LDs (34)(35)(36).…”

“…To study this possibility, the lipin-1-deficient human macrophages were acutely exposed to a high concentration of oleic acid (100 mM), which induces rapid LD formation (34)(35)(36). Inspection of the LDs formed under these conditions revealed notable differences between lipin-1-depleted and control cells (Fig.…”

Featuring… Susumu Mitsutake

“…On the other hand, in adipocytes CERK is involved in the biogenesis of lipid droplets [3], and I doubt it is directly involved in the secretion of MCP1. Macrophage infiltration in the adipose tissue, and the formation of crown like structures, are necessary to stimulate MCP1 secretion in adipocytes.…”