Carrión-Vázquez M, Fernández A M, Chowen J, Nieto-Sampedro M
Neural Plasticity Department, Instituto Cajal (CSIC), Madrid, Spain.
Eur J Neurosci. 1998 Mar;10(3):1153-66. doi: 10.1046/j.1460-9568.1998.00130.x.
Physophilin is an oligomeric protein that binds the synaptic vesicle protein synaptophysin constituting a complex that has been hypothesized to form the exocytotic fusion pore. Microsequencing of several physophilin peptides putatively identified this protein as the Ac39 subunit of the V-ATPase. Ac39 has recently been shown to be present in a synaptosomal complex which, in addition to synaptophysin, includes the bulk of synaptobrevin II, and subunits c and Ac115 of the V0 sector of the V-ATPase. We have cloned physophilin from mouse brain and found a differential region of 12 amino acids when compared with the previously reported sequence of Ac39 from bovine adrenal medulla. RT-PCR cloning from the bovine adrenal medulla demonstrates that sequencing errors occurred in the previous cloning study, and shows that the amino acid sequences of physophilin and Ac39 are completely identical. In situ hybridization in rat brain reveals a largely neuronal distribution of Ac39/physophilin mRNA which spatio-temporally correlates with those of subunit c and synaptophysin. Immunohistochemical analysis shows that Ac39/physophilin is mostly concentrated in the neuropil with a pattern identical to subunit A and very similar to synaptophysin. Double-labelling immunofluorescence shows a complete colocalization of Ac39/physophilin with subunit A and a partial colocalization with synaptophysin in the neuropil. Our findings bring anatomical support for the in vivo occurrence of the synaptophysin-Ac39/physophilin interaction and further suggest a coordinated transcription of V-ATPase and synaptophysin genes. A putative role of Ac39/physophilin in the inactivation of the V-ATPase by disassembly of its V1 sector is also discussed.
亲环蛋白是一种寡聚蛋白，它与突触小泡蛋白突触素结合，构成一种复合物，有人推测该复合物形成了胞吐融合孔。对几种亲环蛋白肽段进行微测序后，初步确定该蛋白为V-ATP酶的Ac39亚基。最近发现Ac39存在于一个突触体复合物中，该复合物除了含有突触素外，还包括大部分的突触结合蛋白II，以及V-ATP酶V0区的c亚基和Ac115亚基。我们从小鼠脑中克隆了亲环蛋白，与之前报道的牛肾上腺髓质Ac39序列相比，发现了一个12个氨基酸的差异区域。从牛肾上腺髓质进行RT-PCR克隆表明，之前的克隆研究中出现了测序错误，并且表明亲环蛋白和Ac39的氨基酸序列完全相同。大鼠脑原位杂交显示Ac39/亲环蛋白mRNA主要分布在神经元中，其时空分布与c亚基和突触素的分布相关。免疫组织化学分析表明，Ac39/亲环蛋白主要集中在神经纤维网中，其模式与A亚基相同，与突触素非常相似。双标记免疫荧光显示，Ac39/亲环蛋白与A亚基在神经纤维网中完全共定位，与突触素部分共定位。我们的研究结果为突触素-Ac39/亲环蛋白相互作用在体内的发生提供了解剖学支持，并进一步表明V-ATP酶和突触素基因的协同转录。还讨论了Ac39/亲环蛋白通过拆卸其V1区使V-ATP酶失活的假定作用。
