1 Recruitment to activated tyrosine kinase growth factor receptors of Grb2 and p21 ras leads to downstream activation of the kinases Raf, MAPK/Erk kinase (Mek) and, subsequently, extracellular signal-regulated kinase (Erk). Activated Erk phosphorylates speci®c serine residues within cytosolic phospholipase A 2 (PLA 2 ), promoting enzyme translocation to membranes and facilitating liberation of arachidonic acid (AA). 2 In the A549 human adenocarcinoma cell line dexamethasone inhibited epidermal growth factor (EGF)-stimulated cytosolic PLA 2 (cPLA 2 ) activation and AA release by blocking the recruitment of Grb2 to the activated EGF receptor (EGF-R) through a glucocorticoid receptor (GR)-dependent (RU486-sensitive), transcription-independent (actinomycin-insensitive), mechanism. 3 The dexamethasone-induced block of Grb2 recruitment was parallelled by changes in phosphorylation status and subcellular localization of lipocortin 1 (LC1) and an increase in the amount of the tyrosine phosphoprotein co-localized with EGF-R. 4 Like dexamethasone, peptides containing E-Q-E-Y-V from the N-terminal domain of LC1 also blocked ligand-induced association of Grb2, p21 ras and Raf. 5 Our results point to an unsuspected rapid eect of glucocorticoids, mediated by occupation of GR but not by changes in gene transcription, which is brought about by competition between LC1 and Grb2 leading to a failure of recruitment o signalling factors to EGF-R

1 We have examined the e ects of 12 glucocorticoids as inhibitors of A549 cell growth. 2 Other than cortisone and prednisone, all the glucocorticoids inhibited cell growth and this was strongly correlated (r=0.91) with inhibition of prostaglandin (PG)E 2 formation. 3 The molecular mechanism by which the active steroids prevented PGE 2 synthesis was examined and three groups were identi®ed. Group A drugs did not inhibit arachidonic acid release but inhibited the induction of COX2. Group B drugs were not able to inhibit the induction of COX2 but inhibited arachidonic acid release through suppression of cPLA 2 activation. Group C drugs were apparently able to bring about both e ects. 4 The inhibitory actions of all steroids was dependent upon glucocorticoid receptor occupation since RU486 reversed their e ects. However, group A acted through the NF-kB pathway to inhibit COX2 as the response was blocked by the inhibitor geldanamycin which prevents dissociation of GR and the e ect was blocked by APDC, the NF-kB inhibitor. On the other hand, the group B drugs were not inhibited by NF-kB inhibitors or geldanamycin but their e ect was abolished by the src inhibitor PP2. Group C drugs depended on both pathways. 5 In terms of PGE 2 generation, there is clear evidence of two entirely separate mechanisms of glucocorticoid action, one of which correlates with NF-kB mediated genomic actions whilst the other, depends upon rapid e ects on a cell signalling system which does not require dissociation of GR. The implications for these ®ndings are discussed.

The Ca(2+)- and phospholipid-binding protein Anx-A1 (annexin 1; lipocortin 1) has been described both as an inhibitor of phospholipase A(2) (PLA(2)) activity and as a mediator of glucocorticoid-regulated cell growth and eicosanoid generation. Here we show that, when compared with Anx-A1(+/+) cells, lung fibroblast cell lines derived from the Anx-A1(-/-) mouse exhibit an altered morphology characterized by a spindle-shaped appearance and an accumulation of intracellular organelles. Unlike their wild-type counterparts, Anx-A1(-/-) cells also overexpress cyclo-oxygenase 2 (COX 2), cytosolic PLA(2) and secretory PLA(2) and in response to fetal calf serum, exhibit an exaggerated release of eicosanoids, which is insensitive to dexamethasone (10(-8)- 10(-6) M) inhibition. Proliferation and serum-induced progression of Anx-A1(+/+) cells from G(0)/G(1) into S phase, and the associated expression of extracellular signal-regulated kinase 2 (ERK2), cyclin-dependent kinase 4 (cdk4) and COX 2, is strongly inhibited by dexamethasone, whereas Anx-A1(-/-) cells are refractory to the drug. Loss of the response to dexamethasone in Anx-A1(-/-) cells occurs against a background of no apparent change in glucocorticoid receptor expression or sensitivity to non-steroidal anti-inflammatory drugs. Taken together, these observations suggest strongly that Anx-A1 functions as an inhibitor of signal-transduction pathways that lead to cell proliferation and may help to explain how glucocorticoids regulate these processes.