Imaging local sphingomyelin-rich domains in the plasma membrane using specific probes and advanced microscopy

• Cytolytic proteins, equinatoxin II and lysenin, bind specifically to sphingomyelin.
• Sphingomyelin-specific probes derived from these proteins are developed.
• State-of-the-art microscopy is introduced to visualize sphingomyelin-rich domains.

Sphingomyelin (SM) is one of the major lipids in the mammalian plasma membrane. Multiple lines of evidence suggest that SM plays at least two functional roles in the cell, as a reservoir of lipid second messengers and as a platform for signaling molecules. To understand the molecular organization and dynamics of the SM-rich membrane domains, new approaches have been developed utilizing newly characterized specific SM-binding probes and state-of-the-art microscopy techniques. The toxic protein from the sea anemone, equinatoxin II, has been characterized as a specific probe for SM. The cytolytic protein from the earthworm, lysenin, has also been used as a SM-specific probe for the analysis of the heterogeneity of SM-rich membrane domains. Recently, using a non-toxic form of lysenin, we showed the spatial and temporal localization of SM in the plasma membrane by confocal and super-resolution microscopy. New microscopy techniques have also been introduced by other groups to help visualize membrane lipid domains. Here we review the most recent studies on imaging the SM-rich domains in biological membranes. This article is part of a Special Issue entitled New Frontiers in Sphingolipid Biology.