McEwen Jason M, Kaplan Joshua M
Department of Molecular Biology, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114, USA.
Mol Biol Cell. 2008 Sep;19(9):3836-46. doi: 10.1091/mbc.e08-02-0160. Epub 2008 Jul 2.
The SM protein UNC-18 has been proposed to regulate several aspects of secretion, including synaptic vesicle docking, priming, and fusion. Here, we show that UNC-18 has a chaperone function in neurons, promoting anterograde transport of the plasma membrane soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein Syntaxin-1. In unc-18 mutants, UNC-64 (Caenorhabditis elegans Syntaxin-1) accumulates in neuronal cell bodies. Colocalization studies and analysis of carbohydrate modifications both suggest that this accumulation occurs in the endoplasmic reticulum. This trafficking defect is specific for UNC-64 Syntaxin-1, because 14 other SNARE proteins and two active zone markers were unaffected. UNC-18 binds to Syntaxin through at least two mechanisms: binding to closed Syntaxin, or to the N terminus of Syntaxin. It is unclear which of these binding modes mediates UNC-18 function in neurons. The chaperone function of UNC-18 was eliminated in double mutants predicted to disrupt both modes of Syntaxin binding, but it was unaffected in single mutants. By contrast, mutations predicted to disrupt UNC-18 binding to the N terminus of Syntaxin caused significant defects in locomotion behavior and responsiveness to cholinesterase inhibitors. Collectively, these results demonstrate the UNC-18 acts as a molecular chaperone for Syntaxin transport in neurons and that the two modes of UNC-18 binding to Syntaxin are involved in different aspects of UNC-18 function.
SM蛋白UNC-18被认为可调节分泌的多个方面，包括突触小泡对接、引发和融合。在此，我们表明UNC-18在神经元中具有伴侣功能，促进质膜可溶性N-乙基马来酰亚胺敏感因子附着蛋白受体（SNARE）蛋白Syntaxin-1的顺行运输。在unc-18突变体中，UNC-64（秀丽隐杆线虫Syntaxin-1）在神经元细胞体中积累。共定位研究和碳水化合物修饰分析均表明这种积累发生在内质网中。这种运输缺陷是Syntaxin-1特异性的，因为其他14种SNARE蛋白和两种活性区标记物未受影响。UNC-18通过至少两种机制与Syntaxin结合：与封闭的Syntaxin结合，或与Syntaxin的N端结合。目前尚不清楚这些结合模式中的哪一种介导了UNC-18在神经元中的功能。UNC-18的伴侣功能在预测会破坏Syntaxin两种结合模式的双突变体中被消除，但在单突变体中未受影响。相比之下，预测会破坏UNC-18与Syntaxin N端结合的突变会导致运动行为和对胆碱酯酶抑制剂反应性的显著缺陷。总体而言，这些结果表明UNC-18在神经元中作为Syntaxin运输的分子伴侣发挥作用，并且UNC-18与Syntaxin的两种结合模式参与了UNC-18功能的不同方面。
