Regulation of leukotriene A(4) hydrolase activity in endothelial cells by phosphorylation

被引:23
作者
Rybina, IV [1 ]
Liu, H [1 ]
Gor, Y [1 ]
Feinmark, SJ [1 ]
机构
[1] COLUMBIA UNIV, DEPT PHARMACOL, COLL PHYS & SURG, NEW YORK, NY 10032 USA
关键词
D O I
10.1074/jbc.272.50.31865
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Endothelial cells contain leukotriene (LT) A(4) hydrolase (LTA-H) as detected by Northern and Western blotting, but several studies have been unable to detect the activity of this enzyme. Since LTA-H could play a hey role in determining what biologically active lipids are generated by activated endothelium during the inflammatory process, we studied possible mechanisms by which this enzyme may be regulated, We find that LTA-H is phosphorylated under basal conditions in human endothelial cells and in this state does not exhibit epoxide hydrolase activity (i.e. conversion of LTA(4) to LTB4). LTA-H purified from endothelial cells is efficiently dephosphorylated by incubation with protein phosphatase-1 in the presence of an LTA-H peptide substrate and not at all in the absence of substrate. Under conditions that lead to dephosphorylation, protein phosphatase-1 activates the epoxide hydrolase activity of LTA-H. Using peptide mapping and site-directed mutagenesis, we have identified serine 415 as the site of phosphorylation of LTA-H by a kinase found in endothelial cell cytosol. In parallel, we have studied a human lung carcinoma cell line that expresses active LTA-H. Although these cells have cytosolic kinases that phosphorylate recombinant LTA-H, they do not target serine 415 and thus do not inhibit LTA-H activity. We believe that LTA-H is regulated in intact cells by a kinase/phosphatase cycle and further that the kinase in endothelial cells specifically recognizes and phosphorylates a regulatory site in the LTA-H.
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页码:31865 / 31871
页数:7
相关论文
共 48 条
[1]  
BOYLE WJ, 1991, METHOD ENZYMOL, V201, P110
[2]   APPLICATION OF AN IMMORTALIZED HUMAN ENDOTHELIAL-CELL LINE TO THE LEUKOCYTE - ENDOTHELIAL ADHERENCE ASSAY [J].
BROWN, KA ;
VORA, A ;
BIGGERSTAFF, J ;
EDGELL, CJS ;
OIKLE, S ;
MAZURE, G ;
TAUB, N ;
MEAGER, A ;
HILL, T ;
WATSON, C ;
DUMONDE, DC .
JOURNAL OF IMMUNOLOGICAL METHODS, 1993, 163 (01) :13-22
[3]   SINGLE-STEP METHOD OF RNA ISOLATION BY ACID GUANIDINIUM THIOCYANATE PHENOL CHLOROFORM EXTRACTION [J].
CHOMCZYNSKI, P ;
SACCHI, N .
ANALYTICAL BIOCHEMISTRY, 1987, 162 (01) :156-159
[4]   HUMAN-ENDOTHELIAL CELLS STIMULATE LEUKOTRIENE SYNTHESIS AND CONVERT GRANULOCYTE RELEASED LEUKOTRIENE-A4 INTO LEUKOTRIENE-B4, LEUKOTRIENE-C4, LEUKOTRIENE-D4 AND LEUKOTRIENE-E4 [J].
CLAESSON, HE ;
HAEGGSTROM, J .
EUROPEAN JOURNAL OF BIOCHEMISTRY, 1988, 173 (01) :93-100
[5]   RELATIONSHIP OF CYCLIC-AMP LEVELS IN LEUKOTRIENE-B4-STIMULATED LEUKOCYTES TO LYSOSOMAL-ENZYME RELEASE AND THE GENERATION OF SUPEROXIDE ANIONS [J].
CLAESSON, HE ;
FEINMARK, SJ .
BIOCHIMICA ET BIOPHYSICA ACTA, 1984, 804 (01) :52-57
[6]   AGGREGATION OF RAT POLYMORPHONUCLEAR LEUKOCYTES INVITRO [J].
CUNNINGHAM, FM ;
SHIPLEY, ME ;
SMITH, MJH .
JOURNAL OF PHARMACY AND PHARMACOLOGY, 1980, 32 (05) :377-380
[7]   LEUKOTRIENES PROMOTE PLASMA LEAKAGE AND LEUKOCYTE ADHESION IN POST-CAPILLARY VENULES - INVIVO EFFECTS WITH RELEVANCE TO THE ACUTE INFLAMMATORY RESPONSE [J].
DAHLEN, SE ;
BJORK, J ;
HEDQVIST, P ;
ARFORS, KE ;
HAMMARSTROM, S ;
LINDGREN, JA ;
SAMUELSSON, B .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES, 1981, 78 (06) :3887-3891
[8]   PERMANENT CELL-LINE EXPRESSING HUMAN FACTOR-VIII-RELATED ANTIGEN ESTABLISHED BY HYBRIDIZATION [J].
EDGELL, CJ ;
MCDONALD, CC ;
GRAHAM, JB .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES, 1983, 80 (12) :3734-3737
[9]  
FEINMARK SJ, 1990, METHOD ENZYMOL, V187, P559
[10]  
FEINMARK SJ, 1986, J BIOL CHEM, V261, P6466