A role for sphingomyelin-rich lipid domains in the accumulation of phosphatidylinositol-4,5-bisphosphate to the cleavage furrow during cytokinesis.
ABSTRACT Cytokinesis is a crucial step in the creation of two daughter cells by the formation and ingression of the cleavage furrow. Here, we show that sphingomyelin (SM), one of the major sphingolipids in mammalian cells, is required for the localization of phosphatidylinositol-4,5-bisphosphate (PIP(2)) to the cleavage furrow during cytokinesis. Real-time observation with a labeled SM-specific protein, lysenin, revealed that SM is concentrated in the outer leaflet of the furrow at the time of cytokinesis. Superresolution fluorescence microscopy analysis indicates a transbilayer colocalization between the SM-rich domains in the outer leaflet and PIP(2)-rich domains in the inner leaflet of the plasma membrane. The depletion of SM disperses PIP(2) and inhibits the recruitment of the small GTPase RhoA to the cleavage furrow, leading to abnormal cytokinesis. These results suggest that the formation of SM-rich domains is required for the accumulation of PIP(2) to the cleavage furrow, which is a prerequisite for the proper translocation of RhoA and the progression of cytokinesis.
Article: Cholesterol-induced fluid membrane domains: a compendium of lipid-raft ternary phase diagrams.[show abstract] [hide abstract]
ABSTRACT: The biophysical underpinning of the lipid-raft concept in cellular membranes is the liquid-ordered phase that is induced by moderately high concentrations of cholesterol. Although the crucial feature is the coexistence of phase-separated fluid domains, direct evidence for this in mixtures of cholesterol with a single lipid is extremely sparse. More extensive evidence comes from ternary mixtures of a high chain-melting lipid and a low chain-melting lipid with cholesterol, including those containing sphingomyelin that are taken to be a raft paradigm. There is, however, not complete agreement between the various phase diagrams and their interpretation. In this review, the different ternary phase diagrams of cholesterol-containing systems are presented in a uniform way, using simple x,y-coordinates to increase accessibility for the non-specialist. It is then possible to appreciate the common features and examine critically the discrepancies and hence what direct biophysical evidence there is that supports the raft concept.Biochimica et Biophysica Acta 09/2009; 1788(10):2114-23. · 4.66 Impact Factor
Article: A novel inhibitor of ceramide trafficking from the endoplasmic reticulum to the site of sphingomyelin synthesis.[show abstract] [hide abstract]
ABSTRACT: Ceramide produced at the endoplasmic reticulum (ER) is transported to the lumen of the Golgi apparatus for conversion to sphingomyelin (SM). N-(3-Hydroxy-1-hydroxymethyl-3-phenylpropyl)dodecanamide (HPA-12) is a novel analog of ceramide. Metabolic labeling experiments showed that HPA-12 inhibits conversion of ceramide to SM, but not to glucosylceramide, in Chinese hamster ovary cells. Cultivation of cells with HPA-12 significantly reduced the content of SM. HPA-12 did not inhibit the activity of SM synthase. The inhibition of SM formation by HPA-12 was abrogated when the Golgi apparatus was made to merge with the ER by brefeldin A. Moreover, HPA-12 inhibited redistribution of a fluorescent analog of ceramide, N-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-pentanoyl)-d-erythro-sphingosine (C(5)-DMB-Cer), from intracellular membranes to the Golgi region. Among four stereoisomers of the drug, (1R,3R)-HPA-12, which resembles natural ceramide stereochemically, was found to be the most active, although (1R,3R)-HPA-12 did not affect ER-to-Golgi trafficking of protein. Interestingly, (1R,3R)-HPA-12 inhibited conversion of ceramide to SM little in mutant cells defective in an ATP- and cytosol-dependent pathway of ceramide transport. These results indicated that (1R,3R)-HPA-12 inhibits ceramide trafficking from the ER to the site of SM synthesis, possibly due to an antagonistic interaction with a ceramide-recognizing factor(s) involved in the ATP- and cytosol-dependent pathway.Journal of Biological Chemistry 12/2001; 276(47):43994-4002. · 4.77 Impact Factor
Article: How principles of domain formation in model membranes may explain ambiguities concerning lipid raft formation in cells.[show abstract] [hide abstract]
ABSTRACT: Sphingolipid and cholesterol-rich liquid ordered lipid domains (lipid rafts) have been studied in both eukaryotic cells and model membranes. However, while the coexistence of ordered and disordered liquid phases can now be easily demonstrated in model membranes, the situation in cell membranes remains ambiguous. Unlike the usual situation in model membranes, under most conditions, cell membranes rich in sphingolipid and cholesterol may have a "granular" organization in which the size of ordered and/or disordered domains is extremely small and domains may be of borderline stability. This review attempts to explain the origin of the divergence between of our understanding of rafts in model membranes and in cells, and how the physical properties of model membranes can help explain many of the ambiguities concerning raft formation and properties in cells. How physical principles of ordered domain formation relate to limitations of detergent insolubility and cholesterol depletion methods used to infer the presence of rafts in cells is also discussed. Possible modifications of these techniques that may increase their reliability are considered. It will be necessary to study model membrane systems more closely approximating cell membranes in order gain a complete understanding of raft properties in cells. Very high concentrations of membrane cholesterol and proteins may explain key physical characteristics of domains in cellular membranes, and are the two of the most obvious factors requiring additional study.Biochimica et Biophysica Acta 01/2006; 1746(3):203-20. · 4.66 Impact Factor