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Protein phosphatase 2A activates the proapoptotic
function of BAD in interleukin- 3–dependent lymphoid cells
by a mechanism requiring 14-3-3 dissociation
by Chi-Wu Chiang, Gregory Harris, Cindy Ellig, Shane C. Masters, Romesh
Subramanian, Shirish Shenolikar, Brian E. Wadzinski, and Elizabeth Yang
Blood
Volume 97(5):1289-1297
March 1, 2001
©2001 by American Society of Hematology
Okadaic acid inhibits BAD dephosphorylation.(A) FL5.12BCL-XL/BAD cells were treated with 0.5
μM OA or dimethyl sulfoxide (DMSO) for 4 hours in the presence (+) or absence (−) of IL-3.
Chi-Wu Chiang et al. Blood 2001;97:1289-1297
©2001 by American Society of Hematology
PP2A-selective inhibitors block BAD dephosphorylation in vivo.(A) FL5.12BCL-XL/BAD cells
were treated with PP1 and PP2A inhibitors calyculin A, fostriecin, and tautomycin in the
presence of IL-3 for 3 hours.
Chi-Wu Chiang et al. Blood 2001;97:1289-1297
©2001 by American Society of Hematology
PP2A activity is significantly inhibited by in vivo OA and fostriecin treatment.(A) Phosphatase
activity in lysates of cells treated with 0.5 μM OA (OA) using either 32P-labeled phosphorylase a
(upper panel) or 32P-labeled histone H1 (lower panel) as substr...
Chi-Wu Chiang et al. Blood 2001;97:1289-1297
©2001 by American Society of Hematology
14-3-3 prevents BAD dephosphorylation in vitro.(A) FL5.12BCL-XL/BAD lysates were prepared in
phosphatase buffer A with or without 1% empigen BB (Emp) (upper panel) or 25 μM R18 peptide
(lower panel).
Chi-Wu Chiang et al. Blood 2001;97:1289-1297
©2001 by American Society of Hematology
PP2A-selective inhibitor, but not PP1-specific inhibitor, inhibits BAD dephosphorylation in
vitro.(A) FL5.12BCL-XL/BAD lysates were prepared in phosphatase buffer A with or without
fostriecin, in the presence or absence of 25 μM R18 peptide, incubated at 30...
Chi-Wu Chiang et al. Blood 2001;97:1289-1297
©2001 by American Society of Hematology
Serine/threonine phosphatase inhibitor calyculin A rescues FL5.12 cells expressing BAD from
apoptosis after IL-3 withdrawal.Cells expressing wild-type BAD (A) or the phosphorylationdefective mutant BAD (B) were treated with 5 nM calyculin A (⋄, ▵) or vehic...
Chi-Wu Chiang et al. Blood 2001;97:1289-1297
©2001 by American Society of Hematology
Activation of the proapoptotic function of BAD by phosphatase in IL-3–dependent FL5.12 cells is
regulated by 14-3-3 binding.In the presence of survival factor IL-3, BAD is phosphorylated and
binds 14-3-3.
Chi-Wu Chiang et al. Blood 2001;97:1289-1297
©2001 by American Society of Hematology