Gangliosides, Glycosyl transferases and newly synthesized gangliosides in Sphingolipid/Cholesterol-enriched domains of Golgi and Plasma membranes from CHO-K1 cells

PILAR CRESPO; ADOLFO ZURITA; CLAUDIO GIRAUDO; HUGO J. MACCIONI; JOSÉ L. DANIOTTI

Bariloche, Río Negro, Argentina

Congreso; XXXIX Reunión Nacional de la Sociedad Argentina de Investigación en Bioquímica y Biología Celular (SAIB); 2003

Sociedad Argentina de Investigación en Bioquímica y Biología Celular (SAIB)

GANGLIOSIDES, GANGLIOSIDE GLYCOSYL TRANSFERASES AND NEWLY SYNTHESIZED GANGLIOSIDES IN SPHINGOLIPID/CHOLESTEROL-ENRICHED DOMAINS OF GOLGI AND PLASMA MEMBRANES FROM CHO-K1 CELLS Pilar Crespo, Adolfo Zurita, Claudio Giraudo, Hugo J. Maccioni and Jose L. Daniotti. CIQUIBIC, Fac. de Ciencias Químicas, UNC, Córdoba. E-mail: daniotti@dqb.fcq.unc.edu.ar Glycosphingolipid-enriched microdomains (GEM) are specialized detergent-resistant domains of the plasma membrane (PM) in which some gangliosides concentrate. Although genesis of GEM is consider to occur in the Golgi complex, where the synthesis of gangliosides also occurs, the issue concerning the incorporation of ganglioside species into GEM is poorly understood. Using CHO-K1 cells with different glycolipid composition, we compared the behaviour to cold Triton X-100 solubilization of PM ganglioside species with the same species newly synthesized in Golgi membranes. We also investigated if three ganglioside glycosyltransferases are included or excluded from GEM in Golgi membranes. Results show that a fraction of PM GM3, and most GD3 and GT3 reside in GEM. Immunocytochemical examination of GD3 expressing cells showed GD3 distributed as cold detergent resistant patches in the PM. In Golgi membranes we were unable to find evidence for GEM localization of either ganglioside glycosyltransferases or newly synthesized gangliosides. Since the same ganglioside species appear in PM GEM, it was concluded that in vivo nascent GD3, GT3 and GM3 segregate from their synthesizing transferases and then enter GEM. This event could have taken place shortly after synthesis in the Golgi cisternae, along the secretory pathway and/or at the cell surface. Supported by Foncyt, HHMI, F. Antorchas, SECyT-UNC, ISN.