Quevedo María F, Lucchesi Ornella, Bustos Matías A, Pocognoni Cristian A, De la Iglesia Paola X, Tomes Claudia N
From the IHEM, Universidad Nacional de Cuyo, CONICET, Facultad de Ciencias Médicas, CC56. 5500 Mendoza, Argentina.
From the IHEM, Universidad Nacional de Cuyo, CONICET, Facultad de Ciencias Médicas, CC56. 5500 Mendoza, Argentina
J Biol Chem. 2016 Oct 28;291(44):23101-23111. doi: 10.1074/jbc.M116.729954. Epub 2016 Sep 9.
At the final stage of exocytotis, a fusion pore opens between the plasma and a secretory vesicle membranes; typically, when the pore dilates the vesicle releases its cargo. Sperm contain a large dense-core secretory granule (the acrosome) whose contents are secreted by regulated exocytosis at fertilization. Minutes after the arrival of the triggering signal, the acrosomal and plasma membranes dock at multiple sites and fusion pores open at the contact points. It is believed that immediately afterward, fusion pores dilate spontaneously. Rab3A is an essential component of human sperm exocytotic machinery. Yet, recombinant, persistently active Rab3A halts calcium-triggered secretion when introduced after docking into streptolysin O-permeabilized cells; so does a Rab3A-22A chimera. Here, we applied functional assays, electron and confocal microscopy to show that the secretion blockage is due to the stabilization of open fusion pores. Other novel findings are that sperm SNAREs engage in α-SNAP/NSF-sensitive complexes at a post-fusion stage. Complexes are disentangled by these chaperons to achieve vesiculation and acrosomal contents release. Thus, post-fusion regulation of the pores determines their expansion and the success of the acrosome reaction.
在胞吐作用的最后阶段，质膜与分泌囊泡膜之间会形成一个融合孔；通常情况下，当孔扩张时，囊泡会释放其内含物。精子含有一个大的致密核心分泌颗粒（顶体），其内容物在受精时通过调节性胞吐作用分泌。触发信号到达几分钟后，顶体膜和质膜在多个位点对接，接触点处的融合孔打开。据信，在此之后，融合孔会自发扩张。Rab3A是人类精子胞吐机制的一个重要组成部分。然而，重组的、持续活跃的Rab3A在对接后引入经链球菌溶血素O通透处理的细胞中时，会阻止钙触发的分泌；Rab3A-22A嵌合体也是如此。在这里，我们应用功能分析、电子显微镜和共聚焦显微镜来表明分泌受阻是由于开放融合孔的稳定。其他新发现是，精子SNARE蛋白在融合后阶段参与α-SNAP/NSF敏感复合物。这些伴侣蛋白会解开复合物以实现囊泡化和顶体内容物释放。因此，融合孔的融合后调节决定了它们的扩张以及顶体反应的成功。
