Ramos R G, Grimwade B G, Wharton K A, Scottgale T N, Artavanis-Tsakonas S
Department of Biology, Yale University, New Haven, Connecticut 06520.
Genetics. 1989 Oct;123(2):337-48. doi: 10.1093/genetics/123.2.337.
Notch is a developmentally regulated locus which controls the differentiation of various Drosophila tissues, among them the embryonic nervous system. Molecular analysis has suggested that Notch is defined by an approximately 40-kb transcription unit which is spliced into a 10.2-kb mRNA composed of nine exonic regions and coding for a 2703-amino acid long transmembrane protein that shows homology to the mammalian epidermal growth factor. Here, we define the 5' end of the transcription unit and determine the sequences deleted in a Notch mutation involving the 5' nontranscribed region. Using a Notch cosmid vector we demonstrate by P element-mediated transformation that all sequences necessary for Notch function are confined in an approximately 40-kb long genomic region. cDNA sequences are used to construct a 15-kb "minigene" which lacks most, but not all, introns and its functionality is also tested by P element transformation. We show that, unlike the cosmid vector which is capable of rescuing completely all Notch mutations, only certain phenotypes can be rescued by the "minigene." The functional implications of our findings are discussed.
Notch是一个受发育调控的基因座，它控制着果蝇各种组织的分化，其中包括胚胎神经系统。分子分析表明，Notch由一个约40kb的转录单元所界定，该转录单元被剪接成一个10.2kb的mRNA，由9个外显子区域组成，编码一个2703个氨基酸长的跨膜蛋白，该蛋白与哺乳动物表皮生长因子具有同源性。在此，我们确定了转录单元的5'端，并确定了涉及5'非转录区域的Notch突变中缺失的序列。使用Notch黏粒载体，我们通过P因子介导的转化证明，Notch功能所需的所有序列都局限于一个约40kb长的基因组区域。cDNA序列用于构建一个15kb的“小基因”，该小基因缺少大部分但并非所有内含子，其功能也通过P因子转化进行测试。我们表明，与能够完全拯救所有Notch突变的黏粒载体不同，“小基因”只能拯救某些表型。我们讨论了这些发现的功能意义。
