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The Pro-apoptotic Function of Death-associated Protein Kinase Is Controlled by a Unique Inhibitory Autophosphorylation-based Mechanism

2001; Elsevier BV; Volume: 276; Issue: 50 Linguagem: Inglês

10.1074/jbc.m105133200

1083-351X

Galit Shohat-Ophir, Taly R. Spivak-Kroizman, Ofer Cohen, Shani Bialik, Gidi Shani, Hanna Berrisi, Miriam Eisenstein, Adi Kimchi,

Endoplasmic Reticulum Stress and Disease

Death-associated protein kinase is a calcium/calmodulin serine/threonine kinase, which positively mediates programmed cell death in a variety of systems. Here we addressed its mode of regulation and identified a mechanism that restrains its apoptotic function in growing cells and enables its activation during cell death. It involves autophosphorylation of Ser 308 within the calmodulin (CaM)-regulatory domain, which occurs at basal state, in the absence of Ca 2+ /CaM, and is inversely correlated with substrate phosphorylation. This type of phosphorylation takes place in growing cells and is strongly reduced upon their exposure to the apoptotic stimulus of C 6 -ceramide. The substitution of Ser 308 to alanine, which mimics the ceramide-induced dephosphorylation at this site, increases Ca 2+ /CaM-independent substrate phosphorylation as well as binding and overall sensitivity of the kinase to CaM. At the cellular level, it strongly enhances the death-promoting activity of the kinase. Conversely, mutation to aspartic acid reduces the binding of the protein to CaM and abrogates almost completely the death-promoting function of the protein. These results are consistent with a molecular model in which phosphorylation on Ser 308 stabilizes a locked conformation of the CaM-regulatory domain within the catalytic cleft and simultaneously also interferes with CaM binding. We propose that this unique mechanism of auto-inhibition evolved to impose a locking device, which keeps death-associated protein kinase silent in healthy cells and ensures its activation only in response to apoptotic signals.