title: Cholesterol depletion disorganizes oocyte membrane rafts altering mouse fertilization.
abstract: Drastic membrane reorganization occurs when mammalian sperm binds to and fuses with the oocyte membrane. Two oocyte protein families are essential for fertilization, tetraspanins and glycosylphosphatidylinositol-anchored proteins. The firsts are associated to tetraspanin-enriched microdomains and the seconds to lipid rafts. Here we report membrane raft involvement in mouse fertilization assessed by cholesterol modulation using methyl-β-cyclodextrin. Cholesterol removal induced: (1) a decrease of the fertilization rate and index; and (2) a delay in the extrusion of the second polar body. Cholesterol repletion recovered the fertilization ability of cholesterol-depleted oocytes, indicating reversibility of these effects. In vivo time-lapse analyses using fluorescent cholesterol permitted to identify the time-point at which the probe is mainly located at the plasma membrane enabling the estimation of the extent of the cholesterol depletion. We confirmed that the mouse oocyte is rich in rafts according to the presence of the raft marker lipid, ganglioside GM1 on the membrane of living oocytes and we identified the coexistence of two types of microdomains, planar rafts and caveolae-like structures, by terms of two differential rafts markers, flotillin-2 and caveolin-1, respectively. Moreover, this is the first report that shows characteristic caveolae-like invaginations in the mouse oocyte identified by electron microscopy. Raft disruption by cholesterol depletion disturbed the subcellular localization of the signal molecule c-Src and the inhibition of Src kinase proteins prevented second polar body extrusion, consistent with a role of Src-related kinases in fertilization via signaling complexes. Our data highlight the functional importance of intact membrane rafts for mouse fertilization and its dependence on cholesterol.
authors: INSERM U1016, Institut Cochin, Université Paris Descartes, 24 rue du Faubourg Saint-Jacques, F75014 Paris, France.
authors: Jorgelina
authors: J
authors: Buschiazzo
authors: Come
authors: C
authors: Ialy-Radio
authors: Jana
authors: J
authors: Auer
authors: Jean-Philippe
authors: JP
authors: Wolf
authors: Catherine
authors: C
authors: Serres
authors: Brigitte
authors: B
authors: Lefèvre
authors: Ahmed
authors: A
authors: Ziyyat
mesh_terms: D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D001896:Boron Compounds / Q000378:metabolism; D051242:Caveolin 1 / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002784:Cholesterol / Q000172:deficiency*; D005260:Female; D005306:Fertilization / Q000187:drug effects; D005677:G(M1) Ganglioside / Q000378:metabolism; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D008297:Male; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D009865:Oocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D010060:Ovulation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000037:antagonists & inhibitors / Q000378:metabolism; D059470:Tetraspanins / Q000378:metabolism
chemical_list: C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D001896:Boron Compounds; D051242:Caveolin 1; D008565:Membrane Proteins; D047428:Protein Kinase Inhibitors; D059470:Tetraspanins; C106354:flotillins; D005677:G(M1) Ganglioside; D002784:Cholesterol; D016392:Proto-Oncogene Proteins pp60(c-src)
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