Ceramide-1-phosphate Binds Group IVA Cytosolic Phospholipase a2 via a Novel Site in the C2 Domain

Stahelin, Robert V.; Subramanian, Preeti; Vora, Mohsin; Cho, Wonhwa; Chalfant, Charles E.

doi:10.1074/jbc.m701396200

Cited by 120 publications

(148 citation statements)

References 49 publications

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“…Cer-1-P interacts with the cationic ␤-groove (Arg-57, Lys-58, and Arg-59) of the C2 domain of cPLA 2 ␣. This enhances interaction with membranes and lowers the EC 50 for Ca 2ϩ down to 31 nM, resulting in increased and catalytic activity in vitro (22,24). Recently, it has been described that CERK is required for cPLA 2 ␣ activation and translocation to the Golgi in response to Ca 2ϩ increases in the cell (25).…”

Section: Discussionmentioning

confidence: 99%

“…Moreover, the single mutation D43N in the C2 domain of the enzyme, which abrogates its Ca 2ϩ binding capacity, does not affect activity in response to FBS. Several reports have established the dispensability of Ca 2ϩ rises in terms of full enzyme activation in vivo (13, 16 -19) while putting forward the requirement of the signaling lipids PtdIns-4,5-P 2 (17,20,21,34) and Cer-1-P to allow interaction with membranes (22)(23)(24)(25). The D43N mutation in cPLA 2 ␣ renders the enzyme refractory to physiological Ca 2ϩ concentrations (31), but it is fully active on PtdIns-4,5-P 2 -containing micelles (34).…”

Section: Discussionmentioning

confidence: 99%

“…However, the increase of intracellular Ca 2ϩ is not universally required, as several reports have shown cPLA 2 ␣ activation in vivo under resting Ca 2ϩ conditions (13, 16 -19). In this regard, anionic phospholipids like phosphatidylinositol 4,5-bisphosphate (PtdIns-4,5-P 2 ) (20, 21) and ceramide 1-phosphate (Cer-1-P) (22)(23)(24)(25) bind specific regions of the enzyme and decrease its Ca 2ϩ requirement to interact with membranes. Also, phosphorylation of cPLA 2 ␣ at Ser residues 505, 515, and 727 appears to play a role in enzyme activation (11,12).…”

Section: Intracellular Lipid Droplets (Ld)mentioning

confidence: 99%

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

Gubern

Barceló-Torns

Barneda

et al. 2009

Journal of Biological Chemistry

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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.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Intracellular Lipid Droplets (Ld)mentioning

confidence: 99%

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

Gubern

Barceló-Torns

Barneda

et al. 2009

Journal of Biological Chemistry

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“…In particular, we have shown that the membrane-associated phosphatidylinositol 4,5-bisphosphate can bind to the "lysine pocket" of GIVA PLA 2 with high affinity and specificity to activate the GIVA PLA 2 independent of Ca 2ϩ (16,17). Another membraneassociated activator, ceramide 1-phosphate, has been shown to bind to the cation groove of the C2 domain to activate GIVA PLA 2 in a Ca 2ϩ -dependent manner (18,19). Calcium binding is crucial for GIVA PLA 2 activation.…”

mentioning

confidence: 99%

Calcium Binding Rigidifies the C2 Domain and the Intradomain Interaction of GIVA Phospholipase A2 as Revealed by Hydrogen/Deuterium Exchange Mass Spectrometry

Hsu

Burke

Stephens

et al. 2008

Journal of Biological Chemistry

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The GIVA phospholipase A 2 (PLA 2 ) contains two domains: a calcium-binding domain (C2) and a catalytic domain. These domains are linked via a flexible tether. GIVA PLA 2 activity is Ca 2؉ -dependent in that calcium binding promotes protein docking to the phospholipid membrane. In addition, the catalytic domain has a lid that covers the active site, presumably regulating GIVA PLA 2 activity. We now present studies that explore the dynamics and conformational changes of this enzyme in solution utilizing peptide amide hydrogen/deuterium (H/D) exchange coupled with liquid chromatographymass spectrometry (DXMS) to probe the solvent accessibility and backbone flexibility of the C2 domain, the catalytic domain, and the intact GIVA PLA 2 . We also analyzed the changes in H/D exchange of the intact GIVA PLA 2 upon Ca 2؉ binding. The DXMS results showed a fast H/D-exchanging lid and a slow exchanging central core. The C2 domain showed two distinct regions: a fast exchanging region facing away from the catalytic domain and a slow exchanging region present in the "cleft" region between the C2 and catalytic domains. The slow exchanging region of the C2 domain is in tight proximity to the catalytic domain. The effects of Ca 2؉ binding on GIVA PLA 2 are localized in the C2 domain and suggest that binding of two distinct Ca 2؉ ions causes tightening up of the regions that surround the anion hole at the tip of the C2 domain. This conformational change may be the initial step in GIVA PLA 2 activation.The cytosolic group IVA (GIVA) 3 phospholipase A 2 (PLA 2 ), also known as cPLA 2 , was purified and cloned in 1991 (1, 2). It is one of the few phospholipases in the phospholipase A 2 superfamily shown to be important in lipid mediator biosynthesis (3, 4). GIVA PLA 2 hydrolyzes membrane phospholipids at the sn-2 position to release free arachidonic acid, which is the precursor of numerous eicosanoids including the prostaglandins and leukotrienes (5, 6) involved in the inflammatory and pain response (7-9). Understanding the regulation of the catalytic activity of GIVA PLA 2 is crucial for understanding eicosanoid metabolism.The activity of the 85-kDa GIVA PLA 2 has been suggested to be regulated by several factors including the intracellular Ca 2ϩ concentration, its phosphorylation state, and the binding of various activators. Since the GIVA PLA 2 was discovered in 1986, the activity of this enzyme has been known to be Ca 2ϩ -dependent (10, 11). The Ca 2ϩ -dependent lipid-binding domain (C2 domain) at the N terminus, which is linked to a catalytic domain where phospholipid hydrolysis occurs, was later identified (12). Further studies of the GIVA PLA 2 activity showed up-regulation by p38 protein kinase mainly through Ser-505 phosphorylation (13-15). Various membrane-associated activators have been shown to bind to GIVA PLA 2 and upregulate its activity. In particular, we have shown that the membrane-associated phosphatidylinositol 4,5-bisphosphate can bind to the "lysine pocket" of GIVA PLA 2 with high affinity and specificity to ac...

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“…ii) On Ca 2+ activation, the C2 domain of cytosolic phospholipase A2 (cPLA2) targets specifically to internal membranes, thereby initiating inflammation (14). Targeting specificity requires the neutral lipid phosphatidylcholine (PC) and the anionic lipid ceramide-1-phosphate (C1P) (10,15). PC binds to charged, polar and hydrophobic side chains near the Ca 2+ site (16,17), whereas C1P binds a separate basic cluster (Arg 2 -Lys) (15).…”

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confidence: 99%

Lipid targeting domain with dual-membrane specificity that expands the diversity of intracellular targeting reactions

Falke

2012

Proc. Natl. Acad. Sci. U.S.A.

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Ceramide-1-phosphate Binds Group IVA Cytosolic Phospholipase a2 via a Novel Site in the C2 Domain

Cited by 120 publications

References 49 publications

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

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

Calcium Binding Rigidifies the C2 Domain and the Intradomain Interaction of GIVA Phospholipase A2 as Revealed by Hydrogen/Deuterium Exchange Mass Spectrometry

Lipid targeting domain with dual-membrane specificity that expands the diversity of intracellular targeting reactions

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