Ara-C- and daunorubicin-induced recruitment of Lyn in sphingomyelinase-enriched membrane rafts.

Abstract

Induction of apoptosis by DNA-damaging agents such as 1-beta-D-arabinofuranosylcytosine (Ara-C) includes the activation of Lyn protein tyrosine kinase. We have previously established that Ara-C-induced activation of Lyn results in its binding to a neutral sphingomyelinase (SMase) and is requisite for its stimulation and the induction of apoptosis in U937 cells. However, the spacio-temporal organization of these events is unclear. This study demonstrates that part of the total cellular SMase activity is sequestered in sphingomyelin-enriched plasma membrane microdomains (rafts). Under Ara-C and daunorubicin (DNR) treatment, Lyn is rapidly activated and translocated into rafts. The compartmentalization of Lyn (as well as neutral SMase activation and apoptosis) induced by these drugs was blocked by the tyrosine kinase inhibitor herbimycin A and raft disruption. In conclusion, this study establishes that DNA-damaging agents such as Ara-C and DNR rapidly induce Lyn activation and its translocation into membrane rafts. This, in turn leads to neutral SMase activation and raft-associated sphingomyelin hydrolysis with the concomitant generation of the proapoptotic lipid second messenger, ceramide. The apparent topological partitioning between DNA damage and apoptosis signaling (integrated into specialized plasma membrane domains) is discussed.