MAPK activation is required for AA release in
IMLF+/+ in response to serum and PMA.
[3H]AA-labeled IMLF+/+ were preincubated with 10
μm SB203580 (15 min), 10 μm U0126 (15 min), 1
μm wortmannin (30 min), 10 μm KN93 (30 min), 10
μm GF109203X (60 min), or 15 ng/μl CHX (30 min) or no
treatment followed by stimulation with serum for 10 min (A, C, and
E) or PMA for 45 min (B, D, and F).
[3H]AA released into the medium was measured and presented as a
percentage of total cellular [3H]AA. G, IMLF+/+
were preincubated with SB203580, U0126, or CHX or no treatment followed by
stimulation with serum for 10 min or PMA for 45 min, as described above. Cell
lysates of unstimulated (US) or stimulated IMLF+/+ were
analyzed by Western blotting to determine activation of ERKs (p-ERK)
or phosphorylation of cPLA2α on Ser-505
(p-cPLA2α) using phosphospecific antibodies. Sample
loading was determined using antibodies to total cPLA2α or
antibodies to β-tubulin. Data represent the average of three experiments
± S.E. (A-D), representative of three independent experiments
in duplicate ± S.D. (E and F) or a representative
Western blot of three independent experiments (G).