Lipid composition of membrane rafts, isolated with and without detergent, from the spleen of a mouse model of Gaucher disease.
Biological membranes are composed of functionally relevant liquid-ordered and liquid-disordered domains that coexist. Within the liquid-ordered domains are low-density microdomains known as rafts with a unique lipid composition that is crucial for their structure and function. Lipid raft composition is altered in sphingolipid storage disorders, and here we determined the lipid composition using a detergent and detergent-free method in spleen tissue, the primary site of pathology, in a mouse model of the sphingolipid storage disorder, Gaucher disease. The accumulating lipid, glucosylceramide, was 30- and 50-fold elevated in the rafts with the detergent and detergent-free method, respectively. Secondary accumulation of di- and trihexosylceramide resided primarily in the rafts with both methods. The phospholipids distributed differently with more than half residing in the rafts with the detergent-free method and less than 10% with the detergent method, with the exception of the fully saturated species that were primarily in the rafts. Individual isoforms of sphingomyelin correlated with detergent-free extraction and more than half resided in the raft fractions. However, this correlation was not seen with the detergent extraction method as sphingomyelin species were spread across both the raft and non-raft domains. Therefore caution must be exercised when interpreting phospholipid distribution in raft domains as it differs considerably depending on the method of isolation. Importantly, both methods revealed the same lipid alterations in the raft domains in the spleen of the Gaucher disease mouse model highlighting that either method is appropriate to determine membrane lipid changes in the diseased state.
Genetics and Molecular Pathology, SA Pathology at Women's and Children's Hospital, 72 King William Road, North Adelaide, South Australia 5006, Australia.
Kathryn J
KJ
Hattersley
Leanne K
LK
Hein
Maria
M
Fuller
D000818:Animals; D002784:Cholesterol / Q000032:analysis; D003902:Detergents / Q000737:chemistry; D004195:Disease Models, Animal; D005776:Gaucher Disease / Q000378:metabolism*; D008055:Lipids / Q000032:analysis*; D021962:Membrane Microdomains / Q000737:chemistry*; D051379:Mice; D018345:Mice, Knockout; D010743:Phospholipids / Q000032:analysis; D013107:Sphingolipids / Q000032:analysis; D013154:Spleen / Q000737:chemistry*; D001617:beta-Glucosidase / Q000235:genetics
D003902:Detergents; D008055:Lipids; D010743:Phospholipids; D013107:Sphingolipids; D002784:Cholesterol; D001617:beta-Glucosidase
BMP; Cer; DHC; Detergent-resistant membranes; GC; GD; Gaucher disease; LC/ESI-MS/MS; Lipid rafts; MBS; MES buffered saline; Membrane microdomains; PC; PE; PG; PI; PS; Phospholipid; SM; Sphingolipid; THC; bis(monoacylglycero)phosphate; ceramide; dihexosylceramide; glucosylceramide; liquid chromatography/electrospray-ionization tandem mass spectrometry; phosphatidylcholine; phosphatidylethanolamine; phosphatidylglycerol; phosphatidylinositol; phosphatidylserine; sphingomyelin; trihexosylceramide