Molecular cloning of a human, hemicholinium-3-sensitive choline transporter
Molecular cloning of a human, hemicholinium-3-sensitive choline transporter
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DOI：
10.1006/bbrc.2000.3561
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发表时间：
2000-10-05
影响因子：
3.1
通讯作者：
Blakely, RD
中科院分区：
文献类型：
--
作者：
Apparsundaram, S;Ferguson, SM;Blakely, RD
在许多生理环境下，胆碱能神经元对Na~+和Cl~-依赖、HC-3敏感、高亲和力的胆碱摄取(HACU)被认为是乙酰胆碱(ACh)生物合成的限速过程。根据人类基因组计划提供的序列信息和最近报道的大鼠CHT1(RCHT1)，我从脊髓中克隆了一个人CHT基因。HCHT基因编码580个氨基酸，与秀丽线虫同源基因CHO-1有93%和51%的同源性，与Na+偶联葡萄糖转运蛋白(SGLT)基因家族成员NaF偶联葡萄糖(SGLT)、核苷和碘转运蛋白有较远的亲缘关系。Northern印迹分析显示，一个类似于5kb转录本的表达在人脑中富含胆碱能神经元的区域，包括壳核、脊髓和延髓。HCHT基因在COS-7细胞中的表达导致细胞膜上可饱和的Na+/Cl-依赖的胆碱摄取(K-m=1.2 mU M)和膜组分中的[H-3]HC-3结合(K-d=4 nM)，这与哺乳动物胆碱能神经元的特征一致。利用辐射杂交作图技术，Me将hCHT基因定位在人类染色体2Q12上。这些研究阐明了hCHT的一级结构和染色体定位，为HACU调节的机制分析和研究hCHT在疾病状态中的作用提供了基础。(C)2000年学术出版社。
Under many physiological circumstances, Na+- and Cl--dependent, hemicholinium-3 (HC-3)-sensitive, high-affinity choline uptake (HACU) in cholinergic neurons is thought to be rate-limiting in the biosynthesis of acetylcholine (ACh). Based on sequence information provided by the Human Genome Project and the recently reported rat CHT1 (rCHT1), me cloned a human CHT cDNA from spinal cord. The hCHT cDNA encodes a protein of 580 amino acids having 93% identity to rCHT1 and 51% identity to the Caenorhabditis elegans homolog CHO-1, and is distantly related to members of the Na+-coupled glucose transporter (SGLT) gene family of Naf-coupled glucose (SGLT), nucleoside and iodide transporters. Northern blot analysis reveals the expression of a similar to 5 kb transcript in human brain regions rich in cholinergic neurons including the putamen, spinal cord, and medulla. Expression of hCHT cDNA in COS-7 cells results in saturable, Na+/Cl--dependent choline uptake (K-m = 1.2 mu M) in membrane vesicles and [H-3] HC-3 binding (K-d = 4 nM) in membrane fractions, consistent with characteristics reported in mammalian cholinergic neurons. Using radiation hybrid mapping techniques, me localized the hCHT gene to human chromosome 2q12. These studies elucidate the primary structure and chromosomal localization of hCHT and provide a basis for mechanistic analysis of HACU regulation and an investigation of the role of hCHT in disease states. (C) 2000 Academic Press.