Nature and subcellular localization of 1H-NMR-detectable mobile lipid domains (ML) were investigated by NMR, Nile red fluorescence and electron microscopy, in NIH-3T3 fibroblasts and their H-ras transformants (3T3ras) transfected with a high number of oncogene copies. Substantial ML levels (ratio of (CH2)n/CH3 peak areas R=1. 56+/-0.33) were associated in untransformed fibroblasts with both (a) intramembrane amorphous lipid vesicles, about 60 nm in diameter, distinct from caveolae; and (b) cytoplasmic, osmiophilic lipid bodies surrounded by own membrane, endowed of intramembrane particles. 2D NMR maps demonstrated that ML comprised both mono- and polyunsaturated fatty chains. Lower ML signals were detected in 3T3ras (R=0.76+/-0.37), under various conditions of cell growth. Very few (if any) lipid bodies and vesicles were detected in the cytoplasmic or membrane compartments of 3T3ras cells with R<0.4, while only intramembrane lipid vesicles were associated with moderate R values. Involvement of phosphatidylcholine hydrolysis in ML generation was demonstrated by selective inhibition of endogenous phospholipase C (PC-plc) or by exposure to bacterial PC-plc. This study indicates that: (1) both cytoplasmic lipid bodies and membrane vesicles (possibly in mutual dynamic exchange) may contribute (although to a different extent) to ML signals; and (2) high levels of ras-transfection either inhibit ML formation or facilitate their extrusion from the cell.

Biophysical and structural characterization of 1H NMR-detectable mobile lipid domains in NIH-3T3 fibroblasts / Ferretti, A.; Knijn, A.; Iorio, E.; Pulciani, S.; Giambenedetti, M.; Molinari, A.; Meschini, S.; Stringaro, A.; Calcabrini, A.; Freitas, I.; Strom, Roberto; Arancia, G; Podo, F.. - In: BIOCHIMICA ET BIOPHYSICA ACTA. - ISSN 0006-3002. - STAMPA. - 1438:(1999), pp. 329-348. [10.1016/S1388-1981(99)00071-2]

Biophysical and structural characterization of 1H NMR-detectable mobile lipid domains in NIH-3T3 fibroblasts.

STROM, Roberto;
1999

Abstract

Nature and subcellular localization of 1H-NMR-detectable mobile lipid domains (ML) were investigated by NMR, Nile red fluorescence and electron microscopy, in NIH-3T3 fibroblasts and their H-ras transformants (3T3ras) transfected with a high number of oncogene copies. Substantial ML levels (ratio of (CH2)n/CH3 peak areas R=1. 56+/-0.33) were associated in untransformed fibroblasts with both (a) intramembrane amorphous lipid vesicles, about 60 nm in diameter, distinct from caveolae; and (b) cytoplasmic, osmiophilic lipid bodies surrounded by own membrane, endowed of intramembrane particles. 2D NMR maps demonstrated that ML comprised both mono- and polyunsaturated fatty chains. Lower ML signals were detected in 3T3ras (R=0.76+/-0.37), under various conditions of cell growth. Very few (if any) lipid bodies and vesicles were detected in the cytoplasmic or membrane compartments of 3T3ras cells with R<0.4, while only intramembrane lipid vesicles were associated with moderate R values. Involvement of phosphatidylcholine hydrolysis in ML generation was demonstrated by selective inhibition of endogenous phospholipase C (PC-plc) or by exposure to bacterial PC-plc. This study indicates that: (1) both cytoplasmic lipid bodies and membrane vesicles (possibly in mutual dynamic exchange) may contribute (although to a different extent) to ML signals; and (2) high levels of ras-transfection either inhibit ML formation or facilitate their extrusion from the cell.
1999
3T3 cells; Electron microscopy; Magnetic Resonance Spectroscopy; Mobile lipid domains; Transformed cell lines
01 Pubblicazione su rivista::01a Articolo in rivista
Biophysical and structural characterization of 1H NMR-detectable mobile lipid domains in NIH-3T3 fibroblasts / Ferretti, A.; Knijn, A.; Iorio, E.; Pulciani, S.; Giambenedetti, M.; Molinari, A.; Meschini, S.; Stringaro, A.; Calcabrini, A.; Freitas, I.; Strom, Roberto; Arancia, G; Podo, F.. - In: BIOCHIMICA ET BIOPHYSICA ACTA. - ISSN 0006-3002. - STAMPA. - 1438:(1999), pp. 329-348. [10.1016/S1388-1981(99)00071-2]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11573/2198
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