After sperm-oocyte fusion, cortical granules (CGs) located in oocyte cortex undergo exocytosis and their content is released into the perivitelline space to avoid polyspermy. Thus, cortical granule exocytosis (CGE) is a key process for fertilization success. We have demonstrated that alpha-SNAP -and its functional partner NSF- mediate fusion of CGs with the plasma membrane in mouse oocytes. Here, we examined at cellular and ultrastructural level oocytes from hyh (hydrocephalus with hop gait) mice, which present a missense mutation in the Napa gene that results in the substitution of methionine for isoleucine at position 105 (M105I) of alpha-SNAP. Mutated alpha-SNAP was mislocalized in hyh oocytes while NSF expression increased during oocyte maturation. Staining of CGs showed that 9.8% of hyh oocytes had abnormal localization of CGs and oval shape. Functional tests showed that CGE was impaired in hyh oocytes. Interestingly, in vitro fertilization assays showed a decreased fertilization rate for hyh oocytes. Furthermore, fertilized hyh oocytes presented an increased polyspermy rate compared to wild type ones. At ultrastructural level, hyh oocytes showed small mitochondria and a striking accumulation and secretion of degradative structures. Our findings demonstrate the negative effects of alpha-SNAP M105 mutation on oocyte biology and further confirm the relevance of alpha-SNAP in female fertility.
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The authors acknowledge the help of Jaime Romaní and Hernán Salinas in the maintenance of the animal facility. MdP is thankful to CONICET, Argentina, for PhD fellowships. MPM is thankful to CONICYT, Chile, for PhD fellowships. This work was supported by Chilean FONDECYT Regular grants 1141015 (LFB), CISNe-UACh (LFB), and DID-UACh (LFB), PLISSER (MdP), and Universidad Nacional de Cuyo SECTYP 06/M93 (MAM).
© 2019, The Author(s).