DAP-kinase participates in TNF-alpha- and Fas-induced apoptosis and its function requires the death domain

Death-associated protein (DAP)-kinase is a calcium/calmodulin regulated ser ine/threoraine 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 antisense RNA protected cells from killing by anti-Fas/APO-1 agonistic antibodies. Deletion of the death domain abrogated the apoptotic functions of the kinase, thus, documenting for the first time the importan ce of this protein domain. Overexpression of a fragment encompassing the de ath domain of DAP-kinase acted as a specific dominant negative mutant that protected cells from TNF-alpha, Fas, and FADD/MORT1-induced death. DAP-kina se apoptotic function was blocked by bcl-2 as well as by crmA and p35 inhib itors of caspases, but not by the dominant negative mutants of FADD/MORT1 o r of caspase 8. Thus, it functions downstream to the receptor complex and u pstream to other caspases. The multidomain structure of this serine/threoni ne kinase, combined with its involvement in cell death induced by several d ifferent triggers, place DAP-kinase at one of the central molecular pathway s leading to apoptosis.