title: BcR-induced apoptosis involves differential regulation of C16 and C24-ceramide formation and sphingolipid-dependent activation of the proteasome.
abstract: In this study, we describe an ordered formation of long- and very long-chain ceramide species in relation to the progression of B-cell receptor (BcR) triggering induced apoptosis. An early and caspase-independent increase in long-chain ceramide species, in which C(16)- ceramide predominated, was observed 6 h after BcR triggering. In contrast, very long-chain ceramide species were generated later, 12-24 h after BcR triggering. The formation of these very long-chain ceramide species, in which C(24)-ceramide predominated, required the activation of effector caspases. BcR-induced formation of long-chain ceramide species resulted in proteasomal activation and degradation of XIAP and subsequent activation of effector caspases, demonstrating an important cell-biological mechanism through which long-chain ceramides may be involved in the progression of BcR triggering induced apoptosis and subsequent formation of very long-chain ceramide species. BcR-induced activation of the proteasome was blocked with ISP-1/myriocin, a potent and selective inhibitor of serine palmitoyl transferase that catalyzes the first and rate-limiting step in the de novo formation of ceramide. Both ISP-1 and clasto-lactacystin beta-lactone, an irreversible inhibitor of the proteasome, prevented BcR cross-linking-induced XIAP degradation. Also, a mutant XIAP lacking the ubiquitin-ligating ring finger motif was completely resistant to proteasome-mediated degradation, and Ramos cells overexpressing XIAP became highly resistant to BcR cross-linking-induced activation of caspases. The formation of C(16)-ceramide in response to BcR cross-linking was found unaltered in XIAP overexpressing Ramos cells, whereas C(24)-ceramide formation was completely abolished. These results demonstrate how de novo generated long-chain ceramide species may be involved in the activation of downstream effector caspases and subsequent formation of very long-chain ceramide species. As such, these results provide novel and important insights into the significance of specific ceramide species in defined stages of apoptosis.
authors.affiliation: Department of Pathology and Laboratory Medicine, Medical Biology branch, University Hospital Groningen, Hanzeplein 1, 9713 GZ Groningen, The Netherlands. b.j.kroesen@thorax.azg.nl
authors.forename: Bart-Jan
authors.initials: BJ
authors.lastname: Kroesen
authors.forename: Susan
authors.initials: S
authors.lastname: Jacobs
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Pettus
authors.forename: Hannie
authors.initials: H
authors.lastname: Sietsma
authors.forename: Jan Willem
authors.initials: JW
authors.lastname: Kok
authors.forename: Yusuf A
authors.initials: YA
authors.lastname: Hannun
authors.forename: Lou F M H
authors.initials: LF
authors.lastname: de Leij
mesh_terms: D017209:Apoptosis; D020169:Caspases; D002518:Ceramides / Q000096:biosynthesis*; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004789:Enzyme Activation; D006801:Humans; D007700:Kinetics; D008928:Mitochondria / Q000502:physiology; D009097:Multienzyme Complexes / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D011506:Proteins / Q000378:metabolism; D011947:Receptors, Antigen, B-Cell / Q000378:metabolism / Q000502:physiology*; D013107:Sphingolipids / Q000502:physiology; D014407:Tumor Cells, Cultured; D051636:X-Linked Inhibitor of Apoptosis Protein
chemical_list: D002518:Ceramides; D009097:Multienzyme Complexes; D011506:Proteins; D011947:Receptors, Antigen, B-Cell; D013107:Sphingolipids; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; C097760:N-palmitoylsphingosine; D020169:Caspases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex