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 longchain 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 ␤-lactone, an irreversible inhibitor of the proteasome, prevented BcR crosslinking-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.

Suppression of sphingomyelin synthase 1 by small interference RNA is associated with enhanced ceramide production and apoptosis after photodamage

“…Similarly, C24-ceramide has been associated with apoptosis [26]. The biological significance of specific fatty acyl chains in ceramides remains to be established.…”

Selective Inhibition of Juxtanuclear Translocation of Protein Kinase C βII by a Negative Feedback Mechanism Involving Ceramide Formed from the Salvage Pathway

“…For example, previous studies have shown that de novo generated ceramide in response to stimulation of the BcR receptor in B cells results in selective accumulation of C 16 -ceramide (23), and this was implicated in the apoptotic response of those cells (24). On the other hand, activation of neutral SMase2 was shown to selectively induce the formation of C 24 -ceramide and C 24:1 -ceramide, and evidence was provided for a role for this pathway in cell cycle regulation (25) and in interleukin-6 formation (26).…”

The BCL-2 Protein BAK Is Required for Long-chain Ceramide Generation during Apoptosis

“…We characterized UV-C-induced ceramide generation in WT BMK cells (Fig. 1c) and found that total ceramide was elevated as early as 2 h following irradiation and continued to increase up to 24 h. Because ceramides with specific chain lengths have been shown to contribute differently to cellular responses to stress stimuli (24,25), we determined the specific ceramides elevated following UV-C irradiation. In WT cells, UV-C irradiation increased most significantly the long-chain ceramides (C 16 , C 18 , and C 20 ; Fig.…”