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Anchoring cortical granules in the cortex ensures trafficking to the plasma membrane for post-fertilization exocytosis.

ABSTRACT: Following fertilization, cortical granules exocytose ovastacin, a metalloendopeptidase that cleaves ZP2 in the zona pellucida surrounding mouse eggs to prevent additional sperm binding. Using high- and super-resolution imaging with ovastacinmCherry as a fluorescent marker, we characterize cortical granule dynamics at single granule resolution in transgenic mouse eggs. Newly-developed imaging protocols provide an unprecedented view of vesicular dynamics near the plasma membrane in mouse eggs. We discover that cortical granule anchoring in the cortex is dependent on maternal MATER and document that myosin IIA is required for biphasic trafficking to the plasma membrane. We observe local clearance of cortical actin during exocytosis and determine that pharmacologic or genetic disruption of trafficking to the plasma membrane impairs secretion of cortical granules and results in polyspermy. Thus, the regulation of cortical granule dynamics at the cortex-plasma membrane interface is critical for exocytosis and the post-fertilization block to sperm binding that ensures monospermic fertilization.

SUBMITTER: Vogt EJ

PROVIDER: S-EPMC6531442 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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Anchoring cortical granules in the cortex ensures trafficking to the plasma membrane for post-fertilization exocytosis.

Vogt Edgar-John EJ Tokuhiro Keizo K Guo Min M Dale Ryan R Yang Guanghui G Shin Seung-Wook SW Movilla Maria Jimenez MJ Shroff Hari H Dean Jurrien J

Nature communications 20190522 1

Following fertilization, cortical granules exocytose ovastacin, a metalloendopeptidase that cleaves ZP2 in the zona pellucida surrounding mouse eggs to prevent additional sperm binding. Using high- and super-resolution imaging with ovastacin<sup>mCherry</sup> as a fluorescent marker, we characterize cortical granule dynamics at single granule resolution in transgenic mouse eggs. Newly-developed imaging protocols provide an unprecedented view of vesicular dynamics near the plasma membrane in mous ...[more]

PMID: 31118423

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Project description:BackgroundOocyte cortical granules are important in the fertilization of numerous species including mammals. Relatively little is known about the composition, migration, and pre-fertilization release of mammalian oocyte cortical granules.ResultsResults obtained with confocal scanning laser microscopy indicated that mouse oocytes have at least two populations of cortical granules, one that bound both the lectin LCA and the antibody ABL2 and one that bound only LCA. Both types of granules were synthesized at the same time during oocyte maturation suggesting that the ABL2 antigen is targeted to specific granules by a sorting sequence. The distribution of both populations of cortical granules was then studied during the germinal vesicle to metaphase II transition. As the oocytes entered metaphase I, the first cortical granule free domain, which was devoid of both populations of cortical granules, formed over the spindle. During first polar body extrusion, a subpopulation of LCA-binding granules became concentrated in the cleavage furrow and underwent exocytosis prior to fertilization. Granules that bound ABL2 were not exocytosed at this time. Much of the LCA-binding exudate from the release at the cleavage furrow was retained in the perivitelline space near the region of exocytosis and was deduced to contain at least three polypeptides with approximate molecular weights of 90, 62, and 56 kDa. A second cortical granule free domain developed following pre-fertilization exocytosis and subsequently continued to increase in area as both, LCA and LCA/ ABL2-binding granules near the spindle became redistributed toward the equator of the oocyte. The pre-fertilization release of cortical granules did not affect binding of sperm to the overlying zona pellucida.ConclusionsOur data show that mouse oocytes contain at least two populations of cortical granules and that a subset of LCA-binding cortical granules is released at a specific time (during extrusion of the first polar body) and place (around the cleavage furrow) prior to fertilization. The observations indicate that the functions of the cortical granules are more complex than previously realized and include events occurring prior to gamete membrane fusion.

Project description:BACKGROUND:Mutations in lysosomal trafficking regulator (LYST) cause Chediak-Higashi syndrome (CHS), a rare immunodeficiency with impaired cytotoxic lymphocyte function, mainly that of natural killer (NK) cells. Our understanding of NK cell function deficiency in patients with CHS and how LYST regulates lytic granule exocytosis is very limited. OBJECTIVE:We sought to delineate cellular defects associated with LYST mutations responsible for the impaired NK cell function seen in patients with CHS. METHODS:We analyzed NK cells from patients with CHS with missense mutations in the LYST ARM/HEAT (armadillo/huntingtin, elongation factor 3, protein phosphatase 2A, and the yeast kinase TOR1) or BEACH (beige and Chediak-Higashi) domains. RESULTS:NK cells from patients with CHS displayed severely reduced cytotoxicity. Mutations in the ARM/HEAT domain led to a reduced number of perforin-containing granules, which were significantly increased in size but able to polarize to the immunologic synapse; however, they were unable to properly fuse with the plasma membrane. Mutations in the BEACH domain resulted in formation of normal or slightly enlarged granules that had markedly impaired polarization to the IS but could be exocytosed on reaching the immunologic synapse. Perforin-containing granules in NK cells from patients with CHS did not acquire certain lysosomal markers (lysosome-associated membrane protein 1/2) but were positive for markers of transport vesicles (cation-independent mannose 6-phosphate receptor), late endosomes (Ras-associated binding protein 27a), and, to some extent, early endosomes (early endosome antigen 1), indicating a lack of integrity in the endolysosomal compartments. NK cells from patients with CHS had normal cytokine compartments and cytokine secretion. CONCLUSION:LYST is involved in regulation of multiple aspects of NK cell lytic activity, ranging from governance of lytic granule size to control of their polarization and exocytosis, as well as regulation of endolysosomal compartment identity. LYST functions in the regulated exocytosis but not in the constitutive secretion pathway.

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Anchoring cortical granules in the cortex ensures trafficking to the plasma membrane for post-fertilization exocytosis.

Publications

Anchoring cortical granules in the cortex ensures trafficking to the plasma membrane for post-fertilization exocytosis.

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