Evidence for segregation of sphingomyelin and cholesterol during formationof COPI-coated vesicles

In higher eukaryotes, phospholipid and cholesterol synthesis occurs mainly in the endoplasmic reticulum, whereas sphingomyelin and higher glycosphingo lipids are synthesized in the Golgi apparatus. Lipids like cholesterol and sphingomyelin are gradually enriched along the secretory pathway, with thei r highest concentration at the plasma membrane. How a cell succeeds in main taining organelle-specific lipid compositions, despite a steady flow of inc oming and outgoing transport carriers along the secretory pathway, is not y et clear. Transport and sorting along the secretory pathway of both protein s and most lipids are thought to be mediated by vesicular transport, with c oat protein I (COPI) vesicles operating in the early secretory pathway. Alt hough the protein constituents of these transport intermediates are charact erized in great detail, much less is known about their lipid content. Using nano-electrospray ionization tandem mass spectrometry for quantitative lip id analysis of COPI-coated vesicles and their parental Golgi membranes, we find only low amounts of sphingomyelin and cholesterol in COPI-coated vesic les compared with their donor Golgi membranes, providing evidence for a sig nificant segregation from COPI vesicles of these lipids. In addition, our d ata indicate a sorting of individual sphingomyelin molecular species. The p ossible molecular mechanisms underlying this segregation, as well as implic ations on COPI function, are discussed.