ROLE OF THE C2B DOMAIN OF SYNAPTOTAGMIN IN VESICULAR RELEASE AND RECYCLING AS DETERMINED BY SPECIFIC ANTIBODY INJECTION INTO THE SQUID GIANT SYNAPSE PRETERMINAL
ROLE OF THE C2B DOMAIN OF SYNAPTOTAGMIN IN VESICULAR RELEASE AND RECYCLING AS DETERMINED BY SPECIFIC ANTIBODY INJECTION INTO THE SQUID GIANT SYNAPSE PRETERMINAL
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DOI：
10.1073/pnas.92.23.10708
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发表时间：
1995-11-07
影响因子：
11.1
通讯作者：
LLINAS, R
中科院分区：
文献类型：
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作者：
FUKUDA, M;MOREIRA, JE;LLINAS, R
Synaptotagmin （Syt）是一种肌醇高聚磷酸系列[IHPS肌醇1,3,4,5-四磷酸肌醇（IP4），肌醇1,3,4,5,6-五磷酸肌醇和肌醇1,2,3,4,4,5,6 -六磷酸肌醇]结合的突触囊泡蛋白。制备了一种针对IHPS结合位点C2B结构域（anti-Syt-C2B）的多克隆抗体。通过比较其抑制IP4与C2B结构域结合的能力和抑制Ca2+/磷脂与C2A结构域结合的能力来确定该抗体对C2B结构域的特异性。在鱿鱼巨突触前注射抗syt - c2b IgG不能阻断突触释放，IP4与抗syt - c2b IgG共注射不能阻断递质的释放，而IP4本身是一种强效的突触释放阻滞剂，注射抗syt - c2b IgG反复刺激突触前纤维，突触后反应幅度迅速下降，可能是由于其阻断突触囊泡循环。电镜显示，注射抗Syt- c2b的突触前突触的突触囊泡数量减少了90%，这些结果表明，Syt分子在Ca2+的突触囊泡融合和IHPS的调节以及突触囊泡的再循环中起着中心作用。
Synaptotagmin (Syt) is an inositol high-polyphosphate series [IHPS inositol 1,3,4,5-tetrakisphosphate (IP4), inositol 1,3,4,5,6-pentakisphosphate, and inositol 1,2,3,4,5,6-hexakisphosphate] binding synaptic vesicle protein. A polyclonal antibody against the C2B domain (anti-Syt-C2B), an IHPS binding site, was produced. The specificity of this antibody to the C2B domain was determined by comparing its ability to inhibit IP4 binding to the C2B domain with that to inhibit the Ca2+/phospholipid binding to the C2A domain. Injection of the anti-Syt-C2B IgG into the squid giant presynapse did not block synaptic release, Coinjection of IP4 and anti-Syt-C2B IgG failed to block transmitter release, while IP4 itself was a powerful synaptic release blocker, Repetitive stimulation to presynaptic fiber injected with anti-Syt-C2B IgG demonstrated a rapid decline of the postsynaptic response amplitude probably due to its block of synaptic vesicle recycling. Electron microscopy of the anti-Syt-C2B-injected presynapse showed a 90% reduction of the numbers of synaptic vesicles, These results, taken together, indicate that the Syt molecule is central, in synaptic vesicle fusion by Ca2+ and its regulation by IHPS, as well as in the recycling of synaptic vesicles.