Nuclear localisation of cytosolic phospholipase A2-alpha in the EA.hy.926 human endothelial cell line is proliferation dependent and modulated by phosphorylation.
Cytosolic phospholipase A(2)-alpha (cPLA(2)-alpha) is a calcium-sensitive enzyme involved in receptor-mediated eicosanoid production. In resting cells, cPLA(2)-alpha is present in the cytosol and nucleus and translocates to membranes via its calcium-dependent lipid-binding (CaLB) domain following stimulation. cPLA(2)-alpha is also regulated by phosphorylation on several residues, which results in enhanced arachidonic acid release. Little is known about the factors controlling the nuclear localisation of cPLA(2)-alpha. Here the nuclear localisation of cPLA(2)-alpha in the EA.hy.926 human endothelial cell line was investigated. Nuclear localisation was dependent on proliferation, with subconfluent cells containing higher levels of nuclear cPLA(2)-alpha than contact-inhibited confluent or serum-starved cells. The broad-range protein kinase inhibitor staurosporine caused a decrease in the nuclear level of cPLA(2)-alpha, whereas the protein phosphatase inhibitor okadaic acid increased the level of nuclear cPLA(2)-alpha. Using inhibitors for specific mitogen-activated protein (MAP) kinases, both p42/44(MAPK) and p38(MAPK) were shown to be important in modulating nuclear localisation. Finally, inhibition of nuclear import and export using Agaricus bisporus lectin and leptomycin B, respectively, demonstrated that cPLA(2)-alpha contains functional nuclear localisation and export signals. Thus we have identified a novel mode of regulation of cPLA(2)-alpha. This, together with the increasing body of evidence supporting the role of nuclear lipid second messengers in gene expression and proliferation, may have important implications for controlling the growth of endothelial cells in angiogenesis and tumour progression.
Grewal
Seema
S
School of Biochemistry and Molecular Biology, University of Leeds, Leeds, LS2 9JT, UK.
Morrison
Ewan E
EE
Ponnambalam
Sreenivasan
S
Walker
John H
JH
D000595:Amino Acid Sequence; D002452:Cell Count; D002455:Cell Division; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000201:enzymology*; D003600:Cytosol / Q000201:enzymology*; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D054513:Group IV Phospholipases A2; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D037102:Lectins / Q000494:pharmacology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019319:Okadaic Acid / Q000494:pharmacology; D010741:Phospholipases A / Q000737:chemistry / Q000378:metabolism*; D054467:Phospholipases A2; D010766:Phosphorylation; D011487:Protein Conformation; D021381:Protein Transport / Q000187:drug effects; D011725:Pyridines / Q000494:pharmacology; D019311:Staurosporine / Q000494:pharmacology
C041433:Agaricus lectins; D005231:Fatty Acids, Unsaturated; D005419:Flavonoids; D007093:Imidazoles; D037102:Lectins; D011725:Pyridines; D019319:Okadaic Acid; D010741:Phospholipases A; D054513:Group IV Phospholipases A2; D054467:Phospholipases A2; D019311:Staurosporine; C093642:SB 203580; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; C038753:leptomycin B