The mixed lineage kinase DLK is oligomerized by tissue transglutaminase during apoptosis

Current evidence suggests that the mixed lineage kinase family member dual leucine zipper-bearing kinase (DLK) might play a significant role in the re gulation of cell growth and differentiation, particularly during the proces s of tissue remodeling. To further explore this working model, we have inve stigated the regulation of host and recombinant DLK in NIH3T3 and COS-1 cel ls undergoing apoptosis, Using calphostin C, a potent and selective inhibit or of protein kinase C and a recognized apoptosis inducer for various cell types, we demonstrate, by immunoblot analysis, that DLK protein levels are rapidly and dramatically down-regulated during the early phases of apoptosi s, Down-regulation in calphostin C-treated cells was also accompanied by th e appearance of SDS- and mercaptoethanol-resistant high molecular weight DL K immunoreactive oligomers, Experiments aimed at elucidating the mechanism( s) underlying DLK oligomerization revealed that the tissue transglutaminase (tTG) inhibitor monodansylcadaverine antagonized the effects of calphostin C almost completely, thereby suggesting the involvement of a tTG-catalyzed reaction as the root cause of DLK downregulation and accumulation as high molecular weight species. In support of this notion, we also show that DLK can serve as a substrate for tTG-dependent cross-linking in vitro and that this covalent post-translational modification leads to the functional inact ivation of DLK, Taken together, these observations suggest that transglutam ination and oligomerization may constitute a relevant physiological mechani sm for the regulation of DLK activity.