Effect of gangliosides on the distribution of a GPI-anchored protein in plasma membrane from CHO-K1 Cells

CRESPO P. M.; ZURITA A. R.; DANIOTTI J.L

Va. Carlos Paz, Córdoba, Argentina

Congreso; XXXVIII Reunión Nacional de la Sociedad Argentina de Investigación en Bioquímica y Biología Celular (SAIB); 2002

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

EFFECTS OF GANGLIOSIDES ON THE DISTRIBUTION OF A GPI-ANCHORED PROTEIN IN PLASMA MEMBRANE. Crespo, P.M., Zurita, A. and   Daniotti, J.L.  CIQUIBIC, Facultad de Ciencias Químicas, U.N.C., E-mail: pcrespo@dqb.fcq.unc.edu.ar   GPI-anchored proteins are mainly clustered in sphingolipid-cholesterol microdomains of the plasma membrane. The distribution of a GPI-anchored fusion protein (GPI-YFP) in the plasma membrane of CHO-K1 cells with different glycolipid compositions was investigated. Cells depleted of glycosphingolipids by inhibiting gluocosylceramide synthase activity, or cell lines expressing different gangliosides due to stable transfection of appropiate ganglioside glycosyltransferases, or exposed to exogenous GM1, were transfected with GPI-YFP cDNA. The distribution of GPI-YFP chimera on the cell surface was investigated by using the membrane-impermeable cross-linking agent bis(sulphosuccinimidyl)suberate (BS3). Results indicate that GPI-YFP form clusters at the surface of cells expressing GM3, or cells depleted of glycolipids, or transfected cells expressing mainly GD3 and GT3, or GM1 and GD1a, or mostly GM2, or highly expressing GM1. However, no significant changes in membrane microdomains of GPI-YFP were detected in the different glycolipid enviroment provided by the membranes of the cells lines under study. On the other hand, wild type CHO-K1 cells loaded with 100µM of GM1 before cross-linking with BS3 showed a dramatic reduction in the efficiency of cross-linking. Results indicate that manipulating the glycolipid content of cellular membrane did not significantly affect the association of GPI-YFP on the cell surface of CHO-K1 cells. The effect excerted by exogenous GM1 gangliosides on GPI-YFP distribution might be a consequence of ganglioside level reached in plasma membrane and/or the effect of particular ganglioside species (micelles) that leads, probably, to membrane architecture and/or dynamic modifications.