Sphingomyelinase-induced phase transformations: causing morphology switches and multiple-time-domain ceramide generation in model raft membranes.

Sphingomyelinase (SMase) has been shown to be involved in a variety of cell regulation processes by reorganizing the cell membrane morphology. Here we report that SMase can induce a reaction-induced and a solvent-mediated phase transformation, causing switches of three stationary membrane morphologies and multiple-time-domain ceramide generation in model raft membranes. The reaction-induced phase transformation, triggered by the addition of SMase, transforms a pre-existing morphology to a long-lasting intermediate morphology with coexisting ceramide-enriched (Cer-enriched) and sphingomyelin-enriched (SM-enriched) domains. Solvent-mediated phase transformation ultimately transforms all of the SM-enriched domains of the intermediate morphology into Cer-enriched domains. Labeled SMase experiments suggest that the intermediate morphology results from physical trapping of SM in the SM-enriched domains, which are found to be relatively inaccessible to SMase. The characterization results from confocal fluorescence imaging show that the trigger of the solvent-mediated phase transformation is the formation of a 3-D feature rich in SMase, sphingomyelin, and ceramide. This 3-D feature is hypothesized as a slowly nucleating SMase-enriched phase, where SMase processes sphingomyelin more efficiently. The disparate time-scales of the formation of these SMase-features and the SM-enriched domains allow for the development of a significant duration of the middle intermediate morphology between the two transformations. The results show that SMase can be actively involved in the lipid membrane phase changes. The multistage morphology evolution is not only due to membrane-compositional changes caused by SMase, but also due to the selective binding of SMase, and the SMase's special phase behavior during the solvent-mediated phase transformation.