Open AccessN‐terminal and C‐terminal plasma membrane anchoring modulate differently agonist‐induced activation of cytosolic phospholipase A 2
Author(s) -
Klapisz Elsa,
Ziari Mouloud,
Wendum Dominique,
Koumanov Kamen,
BrachetDucos Corinne,
Olivier JeanLuc,
Béréziat Gilbert,
Trugnan Germain,
Masliah Joëlle
Publication year - 1999
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1046/j.1432-1327.1999.00797.x
Subject(s) - arachidonic acid , cytosol , phospholipase a2 , biology , microbiology and biotechnology , chinese hamster ovary cell , protein kinase c , extracellular , biochemistry , signal transduction , receptor , enzyme
The 85 kDa cytosolic phospholipase A 2 (cPLA 2 ) plays a key role in liberating arachidonic acid from the sn ‐2 position of membrane phospholipids. When activated by extracellular stimuli, cPLA 2 undergoes calcium‐dependent translocation from cytosol to membrane sites which are still a matter of debate. In order to evaluate the effect of plasma membrane association on cPLA 2 activation, we constructed chimeras of cPLA 2 constitutively targeted to the plasma membrane by the N‐terminal targeting sequence of the protein tyrosine kinase Lck (Lck‐cPLA 2 ) or the C‐terminal targeting signal of K‐Ras4B (cPLA 2 ‐Ras). Constitutive expression of these chimeras in Chinese hamster ovary cells overproducing the α 2B adrenergic receptor (CHO‐2B cells) did not affect the basal release of [ 3 H]arachidonic acid, indicating that constitutive association of cPLA 2 with cellular membranes did not ensure the hydrolysis of membrane phospholipids. However, Lck‐cPLA 2 increased [ 3 H]arachidonic acid release in response to receptor stimulation and to increased intracellular calcium, whereas cPLA 2 ‐Ras inhibited it, compared with parental CHO‐2B cells and CHO‐2B cells producing comparable amounts of recombinant wild‐type cPLA 2 . The lack of stimulation of cPLA 2 ‐Ras was not due to a decreased enzymatic activity as measured using an exogenous substrate, or to a decreased phosphorylation of the protein. These results show that the plasma membrane is a suitable site for cPLA2 activation when orientated correctly.