Laboratoire de biologie des jonctions neuromusculaires normales et pathologiques, Université Paris Descartes, INSERM U686, 45, rue des Saints Pères, 75270 Paris, France.
Chem Biol Interact. 2010 Sep 6;187(1-3):84-9. doi: 10.1016/j.cbi.2010.02.007. Epub 2010 Feb 11.
Normal physiological activity of the neuromuscular junction (NMJ) requires that key molecules are clustered at the synapse. One of these molecules is acetylcholinesterase (AChE) that regulates acetylcholine levels. This enzyme exists under different isoforms but the predominant form at the NMJ is a collagen-tailed enzyme. The collagen associated to AChE (ColQ) fulfills two functions. It anchors and accumulates AChE in the extracellular matrix. Mutations in ColQ lead to faint or no activity of AChE in the synaptic cleft. As a consequence, normal NMJ functioning is impaired and myasthenic syndromes are observed in patients bearing these mutations. Here, we investigated the effects of ColQ deficiency on cholinesterases mRNA levels and cluster formation. We show that overexpression of AChE but not ColQ in muscle cells is sufficient to drive the formation of AChE clusters. The absence of ColQ in muscle cells in vitro and in vivo leads to an increase in AChE(R) and AChE(T) mRNAs, corresponding to two isoforms of AChE. However, AChE activity is decreased in the medium of ColQ-deficient cells suggesting that AChE secretion is impaired. Butyrylcholinesterase (BChE) mRNAs are also upregulated in vivo. Since AChE and BChE can associate with PRiMA, a membrane anchor, we explored the pattern of expression of PRiMA in vitro and in vivo. The level of PRiMA transcripts is downregulated in the absence of ColQ. Therefore, AChE, BChE and PRiMA mRNA level modifications found in the absence of ColQ cannot compensate for the physiological defects observed at the ColQ-deficient NMJs.
神经肌肉接头（NMJ）的正常生理活动需要关键分子在突触处聚集。这些分子之一是乙酰胆碱酯酶（AChE），它调节乙酰胆碱水平。这种酶存在于不同的同工酶中，但在 NMJ 中占主导地位的形式是一种带有胶原蛋白尾巴的酶。与 AChE 相关的胶原蛋白（ColQ）具有两种功能。它将 AChE 锚定并积累在细胞外基质中。ColQ 突变导致突触间隙中 AChE 活性微弱或缺失。因此，正常的 NMJ 功能受损，携带这些突变的患者会出现肌无力综合征。在这里，我们研究了 ColQ 缺乏对胆碱酯酶 mRNA 水平和簇形成的影响。我们表明，肌肉细胞中 AChE 的过表达而不是 ColQ 的过表达足以驱动 AChE 簇的形成。体外和体内肌肉细胞中 ColQ 的缺失导致 AChE(R) 和 AChE(T) mRNA 的增加，对应于 AChE 的两种同工酶。然而，缺乏 ColQ 的细胞中 AChE 活性降低表明 AChE 分泌受损。但酰基胆碱酯酶（BChE）mRNA 也在体内上调。由于 AChE 和 BChE 可以与 PRiMA（一种膜锚）结合，我们探索了 PRiMA 在体外和体内的表达模式。缺乏 ColQ 时，PRiMA 转录本的水平下调。因此，缺乏 ColQ 时发现的 AChE、BChE 和 PRiMA mRNA 水平的修饰不能弥补 ColQ 缺陷 NMJ 观察到的生理缺陷。
