Identification of novel cytosolic phospholipase A(2)s, murine cPLA(2){delta}, {epsilon}, and {zeta}, which form a gene cluster with cPLA(2){beta}.

Phospholipase A(2) hydrolyzes the sn-2 ester bond of glycerophospholipids that produce free fatty acids and lysophospholipids. Cytosolic phospholipase A(2)s (cPLA(2), group IV) are a subgroup of enzymes that act on the intracellular phospholipid membrane. The best investigated cPLA(2)alpha (group IVA) is a key enzyme for lipid mediator production in vivo. Here we report cloning and characterization of novel murine cPLA(2)s: cPLA(2)delta (group IVD), cPLA(2)epsilon (group IVE), and cPLA(2)zeta (group IVF), that form a gene cluster with cPLA(2)beta (group IVB). The deduced amino acid sequences of cPLA(2)delta, epsilon, and zeta demonstrated a conserved domain structure of cPLA(2), i.e. one C2 domain and one lipase domain. The potential catalytic dyad, Ser and Asp, was conserved for these newly cloned cPLA(2)s along with relatively high conservation for the surrounding residues. Transcripts of murine cPLA(2)delta, epsilon, and zeta appeared to be enriched in certain organs rather than ubiquitous distribution. Major Northern signals for cPLA(2)delta were detected in placenta, cPLA(2)epsilon in thyroid, heart, and skeletal muscle, and cPLA(2)zeta in thyroid. Recombinant proteins expressed in human embryonic kidney 293 cells demonstrated molecular sizes of about 100 kDa by Western blotting and exhibited Ca(2+)-dependent PLA(2) activities on 1-palmitoyl-2-[(14)C]arachidonoyl-phosphatidylcholine substrate. In contrast to cPLA(2)alpha, cPLA(2)zeta preferred phosphatidylethanolamine to phosphatidylcholine. Intracellular localization was visualized by green fluorescent-tagged proteins. Each molecule showed specific localization, and cPLA(2)delta translocated from the cytosol to the perinuclear region by calcium-ionophore stimulation. We thus discovered these functional novel cPLA(2) genes, which cluster on murine chromosome 2E5.

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