Membrane restructuring induced by the enzymatic generation of ceramides with very long chain PUFA

PEÑALVA, D.A.; ANTOLLINI, SS.; AMBROGGIO, E.E.; AVELDAÑO, M.I.; FANANI, M.L.

Córdoba

Congreso; Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB); 2016

Spermatozoa are rich in sphingolipids with very long chain PUFA (V). The sphingomyelin (SM) located on the rat sperm head, which almost exclusively contains V with C28-C32 carbon atoms (V-SM), is converted to the corresponding ceramides (Cer) as a result of the acrosomal reaction (AR). Here, the effect of the SM → Cer conversion on the integrity and structuring of membrane bilayers was studied in unilamellar vesicles (LUVs, GUVs) made of POPC/SM binary mixtures. The SM was V-SM or a palmitate-rich SM (P-SM), and the reaction was catalyzed by a SMase. In LUVs of uniform size, Cer generation induced membrane interaction, as indicated by the vesicle aggregation detected by dynamic light scattering, and confirmed by an increase of FRET signal in assays using NBD-PE as donor and Rh-PE as acceptor. The in-plane lateral organization on the vesicles also differed, as shown by fluorescent probes (Laurdan, DPH). Notable disparities were seen if V-SM or P-SM were the SMase substrates. Fluorescence confocal microscopy of SMase-exposed GUVs containing each SM revealed notable contrasts in leakage rate and membrane reorganization. The production of P-Cer resulted in condensed domains formed in unbroken vesicles, whereas that of V-Cer induced an intense bilayer destabilization that led to vesicle collapse. This may respond for part of the membrane events that occur on the sperm head during the AR.