Varoqui H, Erickson J D
Neuroscience Center, Louisiana State University Medical School, New Orleans 70112, USA.
J Physiol Paris. 1998 Apr;92(2):141-4. doi: 10.1016/S0928-4257(98)80152-6.
Chimeras between the human vesicular acetylcholine transporter (hVAChT) and the neuronal isoform of the human vesicular monoamine transporter (hVMAT2) have been constructed and stably expressed in a rat pheochromocytoma cell line (PC12) in an effort to identify cholinergic-specific domains of VAChT. Examination of the transport properties of a chimera in which the N-terminal portion (up to putative transmembrane domain II and including the lumenal glycosylated loop) of hVAChT was replaced with hVMAT2 sequences (2/V@NheI) revealed that its apparent affinity for acetylcholine (ACh) was reduced approximately seven-fold compared to wild-type. However, the affinity of this chimera for vesamicol did not significantly differ from hVAChT. Similarly, the 2/V@NheI chimera retained its preferential targeting to the small synaptic-like vesicles found in PC12 cells in agreement with our recently reported observations that the synaptic vesicle targeting domain resides in the cytoplasmic tail of VAChT.
构建了人囊泡乙酰胆碱转运体（hVAChT）与人囊泡单胺转运体神经元亚型（hVMAT2）之间的嵌合体，并在大鼠嗜铬细胞瘤细胞系（PC12）中稳定表达，以确定VAChT的胆碱能特异性结构域。对一种嵌合体（其中hVAChT的N端部分（直至假定的跨膜结构域II并包括腔内糖基化环）被hVMAT2序列取代（2/V@NheI））的转运特性进行研究，结果显示，与野生型相比，其对乙酰胆碱（ACh）的表观亲和力降低了约7倍。然而，这种嵌合体对vesamicol的亲和力与hVAChT没有显著差异。同样，2/V@NheI嵌合体保留了其对PC12细胞中发现的小突触样囊泡的优先靶向性，这与我们最近报道的观察结果一致，即突触囊泡靶向结构域位于VAChT的细胞质尾部。
