Dap-Kinase Participates in TNF-{alpha}-And FAS-Induced Apoptosis and Its Function Requires the Death Domain

doi:10.1083/jcb.146.1.141

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© The Rockefeller University Press, 0021-9525/1999//141 $5.00
The Journal of Cell Biology, Volume 146, Number 1, , 1999 141-148

Original Article

Dap-Kinase Participates in TNF- ${alpha}$ –And FAS-Induced Apoptosis and Its Function Requires the Death Domain

Ofer Cohen^a, Boaz Inbal^a, Joseph L. Kissil^a, Tal Raveh^a, Hanna Berissi^a, Taly Spivak-Kroizaman^a, Elena Feinstein^a, and Adi Kimchi^a

^a Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel
Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.972-8-9344108972-8-9342428

lvkimchi{at}weizmann.weizmann.ac.il

Death-associated protein (DAP)–kinase is a calcium/calmodulinregulated serine/threonine kinase that carries ankyrin repeats,a death domain, and is localized to the cytoskeleton. Here,we report that this kinase is involved in tumor necrosis factor(TNF)- ${alpha}$ and Fas-induced apoptosis. Expression of DAP-kinase antisenseRNA protected cells from killing by anti–Fas/APO-1 agonisticantibodies. Deletion of the death domain abrogated the apoptoticfunctions of the kinase, thus, documenting for the first timethe importance of this protein domain. Overexpression of a fragmentencompassing the death domain of DAP-kinase acted as a specificdominant negative mutant that protected cells from TNF- ${alpha}$ , Fas,and FADD/MORT1–induced cell death. DAP-kinase apoptoticfunction was blocked by bcl-2 as well as by crmA and p35 inhibitorsof caspases, but not by the dominant negative mutants of FADD/MORT1or of caspase 8. Thus, it functions downstream to the receptorcomplex and upstream to other caspases. The multidomain structureof this serine/threonine kinase, combined with its involvementin cell death induced by several different triggers, place DAP-kinaseat one of the central molecular pathways leading to apoptosis.

Key Words: DAP-kinase • tumor necrosis factor- ${alpha}$ • Fas • death domain • apoptosis

1.used in this paper: ${Delta}$ CAM, deletion of calmodulin regulatorydomain; CARD, caspase-recruiting domain; DAP, death-associatedprotein; DD, death domain; DISC, death-inducing signaling complex;DN-MORT/FADD, dominant negative mutant of MORT/FADD; GFP, greenfluorescent protein; ICE, interleukin 1β–convertingenzyme; IFN- ${gamma}$ , interferon- ${gamma}$ ; PARP,poly (ADP–ribose) polymerase;TNF, tumor necrosis factor; TNF-R, TNF- ${alpha}$ receptor

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