INIBIOLP   05426
INSTITUTO DE INVESTIGACIONES BIOQUIMICAS DE LA PLATA "PROF. DR. RODOLFO R. BRENNER"
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Coupling between ordered and disordered phases in asymmetric lipid bilayers studied by AFM-Force spectroscopy
Autor/es:
ERASMO OVALLE-GARCÍA; IVÁN ORTEGA-BLAKE; ROMINA F. VAZQUEZ; LAURA S. BAKÁS; SABINA M. MATÉ; ARMANDO ANTILLÓN; CARLOS MUÑOZ-GARAY
Lugar:
Michoacán, México
Reunión:
Congreso; XXII Congreso de la rama de Bioenergetica y Biomembranas; 2021
Resumen:
Cell plasma membranes have an asymmetric distribution of phospholipids with sphingolipids present almost exclusively in the outer leaflet of the bilayer, with consequences for the formation of lipid domains across the membrane. In this study, we explored domain formation when palmitoylsphingomyelin (SM) is incorporated into the outer leaflet of dioleoylphosphatidylcholine (DOPC)/cholesterol (Chol) supported bilayers through cyclodextrin (CD)-mediated lipid exchange, producing lipid asymmetry.Phase segregation was assessed by AFM microscopy for different incubation conditions with CD-SM. We found that treatment with 5 mM CD-SM for 30 min produced liquid-ordered (Lo) domains when incubated at 37°C while more ordered domains were observed if the incubation was performed at 24°C. Less defined structures appeared with shorter times while high concentrations led to an almost continuous ordered phase in the outer leaflet.In Force spectroscopy measurements, the force curves acquired in domains with Lo characteristics in the asymmetric bilayers showed two rupture events attributed to a liquid-disordered (Ld) proximal hemilayer that collapsed first at lower applied forces (~7.5 nN) and a Lo outer hemilayer that was punctured by the tip in a second rupture event at higher forces (~13 nN). The rupture depths of each event agreed with these considerations. The continuous Ld phase, on the other hand, showed curves with single rupture events at ~5.6 nN. The higher rupture force registered for the proximal Ld hemilayer in the asymmetric (Lo/Ld) domains (Ld~7.5 nN) compared to the continuous Ld phase having two opposing Ld leaflets (Ld~5.6nN) reflected a degree of interleaflet coupling when the ordered phase was present in the outer hemilayer. In addition, the mechanical properties (rupture force and Young?s modulus) of Lo domains in asymmetric bilayers were similar to that of domains with two opposing Lo leaflets as measured in symmetric DOPC/SM/Chol bilayers.These results support the idea that formation of sphingolipid-enriched ordered domains in the outer hemilayer can modify the physical properties of the inner leaflet of the plasma membrane. These coupling phenomena could play a relevant role in signal transduction across the membrane.