The Drosophila nitric-oxide synthase gene (dNOS) encodes a family of proteins that can modulate NOS activity by acting as dominant negative regulators
The Drosophila nitric-oxide synthase gene (dNOS) encodes a family of proteins that can modulate NOS activity by acting as dominant negative regulators
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DOI：
10.1074/jbc.m105066200
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发表时间：
2001-11-09
影响因子：
4.8
通讯作者：
Enikolopov, G
中科院分区：
文献类型：
--
作者：
Stasiv, Y;Regulski, M;Enikolopov, G
一氧化氮(NO)参与果蝇器官发育、突触形成和对低氧的反应。我们克隆并分析了果蝇基因组中唯一编码果蝇一氧化氮合酶(DNOS)的基因。它由19个外显子组成，分布在34kb的基因组DNA上。选择性转录起始点和选择性剪接位点被用来从DNOS基因中产生各种各样的mRNAs。我们确定了8个新的转录本，它们在整个果蝇发育过程中广泛表达，并编码了一个与DNOS相关的蛋白家族。选择性剪接影响DNOS初级转录本的5‘-非翻译区和编码区。大多数基因编码区的剪接改变会导致开放阅读框的提前终止。因此，没有一个可供选择的转录本编码具有酶活性的蛋白质。然而，其中一些较短的DNOS蛋白产物在哺乳动物细胞中共表达时，可以有效地抑制全长DNOS1蛋白的酶活性，从而成为NO合成的主要负调控因子。利用免疫沉淀，我们证明了这些短的DNOS蛋白亚型可以与DNOS1形成异二聚体，指出了显性负面效应的物理基础。我们的结果表明，果蝇一氧化氮合酶基因编码的蛋白家族具有新的调节功能。
Nitric oxide (NO) is involved in organ development, synaptogenesis, and response to hypoxia in Drosophila. We cloned and analyzed the only gene in the fly genome that encodes Drosophila nitric-oxide synthase (dNOS). It consists of 19 exons and is dispersed over 34 kilobases of genomic DNA. Alternative transcription start sites and alternative splice sites are used to generate a remarkable variety of mRNAs from the dNOS gene. We identified eight new transcripts that are widely expressed throughout Drosophila development and encode a family of DNOS-related proteins. Alternative splicing affects both the 5'-untranslated region and the coding region of the dNOS primary transcript. Most of the splicing alterations in the coding region of the gene lead to premature termination of the open reading frame. As a result, none of the alternative transcripts encode an enzymatically active protein. However, some of these shorter DNOS protein products can effectively inhibit enzymatic activity of the full-length DNOS1 protein when co-expressed in mammalian cells, thus acting as dominant negative regulators of NO synthesis. Using immunoprecipitation, we demonstrate that these short DNOS protein isoforms can form heterodimers with DNOS1, pointing to a physical basis for the dominant negative effect. Our results suggest a novel regulatory function for the family of proteins encoded by the Drosophila NOS gene.