Gengyo-Ando K, Kitayama H, Mukaida M, Ikawa Y
Laboratory of Molecular Oncology, The Institute of Physical and Chemical Research (RIKEN), Tsukuba, Ibaraki 305, Japan.
J Neurosci. 1996 Nov 1;16(21):6695-702. doi: 10.1523/JNEUROSCI.16-21-06695.1996.
Caenorhabditis elegans UNC-18 protein, homologous to yeast Sec1p, is important in neurotransmitter release, because the unc-18 mutation leads to severe paralysis and presynaptic acetylcholine (ACh) accumulation. To examine the functional conservation in mammals, we tried to isolate unc-18 isoforms from mouse and human brain cDNA libraries and obtained two classes of isoforms-neural genes and ubiquitous genes. Neural genes were identical to Munc-18 (also known as n-Sec1 or rbSec1), identified in rat and bovine brains as a syntaxin-binding protein. According to "Munc-18" terminology, we call the neural genes Munc-18-1 and the ubiquitous genes Munc-18-3. These mammalian isoforms exhibit 58% (Munc-18-1) and 42-43% (Munc-18-3) amino acid sequence identity with UNC-18. Next, we constructed transgenic unc-18 mutants to test biological activity of mouse Munc-18-1 and Munc-18-3 under the control of C. elegans unc-18 promoter. Munc-18-1 compensates for severe locomotion disability and cholinergic defects, e.g., abnormal sensitivities to cholinesterase inhibitors and cholinergic receptor agonists in unc-18 mutants, but Munc-18-3 fails. These data suggest that Munc-18-1 and C. elegans unc-18 may play positive roles in ACh release and that the molecular mechanism of neuronal regulated secretion has been partially conserved from nematodes to mammals.
秀丽隐杆线虫的UNC-18蛋白与酵母Sec1p同源，在神经递质释放中起重要作用，因为unc-18突变会导致严重瘫痪和突触前乙酰胆碱（ACh）积累。为了研究哺乳动物中的功能保守性，我们试图从小鼠和人类脑cDNA文库中分离unc-18亚型，获得了两类亚型——神经基因和普遍存在的基因。神经基因与Munc-18（也称为n-Sec1或rbSec1）相同，在大鼠和牛脑中被鉴定为一种与 syntaxin结合的蛋白。根据“Munc-18”术语，我们将神经基因称为Munc-18-1，将普遍存在的基因称为Munc-18-3。这些哺乳动物亚型与UNC-18的氨基酸序列同一性分别为58%（Munc-18-1）和42-43%（Munc-18-3）。接下来，我们构建了转基因unc-18突变体，以测试秀丽隐杆线虫unc-18启动子控制下小鼠Munc-18-1和Munc-18-3的生物活性。Munc-18-1可补偿严重的运动障碍和胆碱能缺陷，例如unc-18突变体对胆碱酯酶抑制剂和胆碱能受体激动剂的异常敏感性，但Munc-18-3则不能。这些数据表明，Munc-18-1和秀丽隐杆线虫unc-18可能在ACh释放中起积极作用，并且神经元调节分泌的分子机制已从线虫到哺乳动物得到部分保守。
