T-CELL APOPTOSIS INDUCED BY INTERLEUKIN-2 DEPRIVATION OR TRANSFORMING GROWTH FACTOR-BETA(2) - MODULATION BY THE PHOSPHATASE INHIBITORS OKADAIC ACID AND CALYCULIN-A

The phosphatase inhibitors okadaic acid and calyculin A were used to examine the role of phosphorylation processes in T cell apoptosis induced by interleukin-2 (IL-2) deprivation or transforming growth factor-beta(2)(TGF-beta(2)). Okadaic acid and calyculin A inhibited IL-2-driven T cell proliferation and induced apoptosis at concentrations known to inhibit protein phosphatase 1, High concentrations of both agents caused toxic changes of prominent cellular swelling and dilatation of rough endoplasmic reticular profiles. When the T cells were induced to undergo apoptosis by IL-2 deprivation, okadaic acid and calyculin A delayed loss of membrane integrity, nucleosomal size DNA fragmentation, and loss of bcl-2 mRNA. However, T cells deprived of IL-2 in the presence of okadaic acid or calyculin A revealed DNA breaks by in situ DNA end labeling and apoptotic morphology by electron microscopy and failed to show enhanced survival after reexposure to IL-2. Although TGF-beta-mediated signaling is thought to involve the dephosphorylation of specific substrates, okadaic acid and calyculin A not only failed to inhibit, but actually augmented, TGF-beta(2)-induced inhibition of T cell proliferation and induction of apoptosis. Exposure to either TGF-beta(2) or the phosphatase inhibitors prevented phosphorylation of the retinoblastoma protein RE. In summary, okadaic acid and calyculin A: (i) induce T cell apoptosis in the presence of IL-2, (ii) allow us to distinguish essential from epiphenomenal features of T cell. apoptosis after IL-2 deprivation, and (iii) cooperate with TGF-beta(2) in inducing growth arrest and apoptosis of murine T cells via intracellular cascades that converge in the prevention of RB phosphorylation. (C) 1995 Academic Press, Inc.