Nicotinamide mononucleotide adenylyl transferase-mediated axonal protection requires enzymatic activity but not increased levels of neuronal nicotinamide adenine dinucleotide.
Nicotinamide mononucleotide adenylyl transferase-mediated axonal protection requires enzymatic activity but not increased levels of neuronal nicotinamide adenine dinucleotide.
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DOI：
10.1523/jneurosci.5469-08.2009
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发表时间：
2009-04-29
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影响因子：
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通讯作者：
Milbrandt J
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文献类型：
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作者：
Sasaki Y;Vohra BP;Lund FE;Milbrandt J
轴突变性是许多神经系统疾病的标志。在神经退行性疾病动物模型中的研究表明，轴突变性是疾病过程中的早期事件，延迟该过程可导致疾病进展减缓和生存期延长。Wallerian变性缓慢（Wlds）蛋白的过表达可以延迟通过轴突切开术、化疗剂或基因突变引发的轴突变性。Wlds蛋白由泛素化因子Ube4b的N末端部分与烟酰胺腺嘌呤二核苷酸（NAD+）生物合成酶烟酰胺单核苷酸腺苷酰转移酶1（Nmnat 1）融合组成。我们以前表明，该融合蛋白的Nmnat 1部分是WLDS介导的轴突保护的关键部分。在这里，我们描述了一个自动化的定量分析评估轴突变性的发展。该方法成功地表明，Nmnat 1酶活性对于轴突保护是重要的，因为酶活性降低的突变体缺乏轴突保护活性。我们还发现，Nmnat酶具有不同的序列和结构，从不同的物种，包括果蝇，酿酒酵母，和古细菌甲烷醛球菌詹氏，它编码的蛋白质没有同源性真核Nmnat酶，所有介导强大的轴突保护轴突切断后。除了Nmnat酶活性的重要性之外，我们没有观察到稳态NAD+水平的变化，并且我们发现抑制哺乳动物细胞中合成Nmnat底物的烟酰胺磷酸核糖基转移酶（Nampt）并不影响Nmnat1的保护活性。这些结果提供了新的Nmnat酶活性在轴突保护中的作用的可能性，除了NAD+的合成。
Axonal degeneration is a hallmark of many neurological disorders. Studies in animal models of neurodegenerative diseases indicate that axonal degeneration is an early event in the disease process, and delaying this process can lead to decreased progression of the disease and survival extension. Overexpression of the Wallerian degeneration slow (Wlds) protein can delay axonal degeneration initiated via axotomy, chemotherapeutic agents, or genetic mutations. The Wlds protein consists of the N-terminal portion of the ubiquitination factor Ube4b fused to the nicotinamide adenine dinucleotide (NAD+) biosynthetic enzyme nicotinamide mononucleotide adenylyl transferase 1 (Nmnat1). We previously showed that the Nmnat1 portion of this fusion protein was the critical moiety for Wlds-mediated axonal protection. Here, we describe the development of an automated quantitative assay for assessing axonal degeneration. This method successfully showed that Nmnat1 enzymatic activity is important for axonal protection as mutants with reduced enzymatic activity lacked axon protective activity. We also found that Nmnat enzymes with diverse sequences and structures from various species, including Drosophila melanogaster, Saccharomyces cerevisiae, and archaebacterium Methanocaldococcus jannaschii, which encodes a protein with no homology to eukaryotic Nmnat enzymes, all mediate robust axonal protection after axotomy. Besides the importance of Nmnat enzymatic activity, we did not observe changes in the steady-state NAD+ level, and we found that inhibition of nicotinamide phosphoribosyltransferase (Nampt), which synthesizes substrate for Nmnat in mammalian cells, did not affect the protective activity of Nmnat1. These results provide the possibility of a role for new Nmnat enzymatic activity in axonal protection in addition to NAD+ synthesis.