Dynamics of SNARE assembly and disassembly during sperm acrosomal exocytosis.
Dynamics of SNARE assembly and disassembly during sperm acrosomal exocytosis.
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DOI：
10.1371/journal.pbio.0030323
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发表时间：
2005-10
期刊：
影响因子：
9.8
通讯作者：
Mayorga LS
中科院分区：
文献类型：
--
作者：
De Blas GA;Roggero CM;Tomes CN;Mayorga LS
膜识别和融合过程中SNARs的组装和分解动力学是细胞内运输和调节分泌的核心问题。精子单个囊泡(顶体)的胞吐作用是一个同步的、要么全有要么全无的过程，在细胞的一生中只发生一次，取决于GTP结合蛋白Rab3和神经毒素敏感SNARs的激活。这些特性使顶体胞吐作用成为研究膜融合级联不同阶段的独特哺乳动物模型。通过使用功能分析和免疫荧光技术，结合神经毒素和光敏的钙离子螯合剂，我们表明，在未激活的精子中，圈套被锁定在异三聚体顺式复合体中。当钙离子进入细胞质后，Rab3被激活，并触发依赖于Nsf/α-SNAP的顺式SNARE复合体的解体。然后，质膜和顶体外膜上的单体陷阱可以在松散的反式复合体中自由重组，这些复合体对Nsf/α-SNAP具有抵抗力，并对两种囊泡相关膜蛋白特异性神经毒素的切割具有不同的敏感性。钙离子必须从顶体内部释放，才能触发膜融合的最后步骤，这需要完全组装的反式SNARE复合体和突触凝集素。我们的结果表明，SNARE复合体的单向和顺序的拆解和组装驱动顶体胞吐。SNARE复合体的单向和顺序拆解和组装驱动精子顶体胞吐。
The dynamics of SNARE assembly and disassembly during membrane recognition and fusion is a central issue in intracellular trafficking and regulated secretion. Exocytosis of sperm's single vesicle—the acrosome—is a synchronized, all-or-nothing process that happens only once in the life of the cell and depends on activation of both the GTP-binding protein Rab3 and of neurotoxin-sensitive SNAREs. These characteristics make acrosomal exocytosis a unique mammalian model for the study of the different phases of the membrane fusion cascade. By using a functional assay and immunofluorescence techniques in combination with neurotoxins and a photosensitive Ca2+ chelator we show that, in unactivated sperm, SNAREs are locked in heterotrimeric cis complexes. Upon Ca2+ entry into the cytoplasm, Rab3 is activated and triggers NSF/α-SNAP-dependent disassembly of cis SNARE complexes. Monomeric SNAREs in the plasma membrane and the outer acrosomal membrane are then free to reassemble in loose trans complexes that are resistant to NSF/α-SNAP and differentially sensitive to cleavage by two vesicle-associated membrane protein (VAMP)–specific neurotoxins. Ca2+ must be released from inside the acrosome to trigger the final steps of membrane fusion that require fully assembled trans SNARE complexes and synaptotagmin. Our results indicate that the unidirectional and sequential disassembly and assembly of SNARE complexes drive acrosomal exocytosis. Unidirectional and sequential disassembly and assembly of SNARE complexes drive sperm acrosomal exocytosis.
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