Gruenbaum Y, Landesman Y, Drees B, Bare J W, Saumweber H, Paddy M R, Sedat J W, Smith D E, Benton B M, Fisher P A
Department of Biochemistry and Biophysics, University of California at San Francisco 94143.
J Cell Biol. 1988 Mar;106(3):585-96. doi: 10.1083/jcb.106.3.585.
A cDNA clone encoding a portion of Drosophila nuclear lamins Dm1 and Dm2 has been identified by screening a lambda-gt11 cDNA expression library using Drosophila lamin-specific monoclonal antibodies. Two different developmentally regulated mRNA species were identified by Northern blot analysis using the initial cDNA as a probe, and full-length cDNA clones, apparently corresponding to each message, have been isolated. In vitro transcription of both full-length cDNA clones in a pT7 transcription vector followed by in vitro translation in wheat germ lysate suggests that both clones encode lamin Dm0, the polypeptide precursor of lamins Dm1 and Dm2. Nucleotide sequence analyses confirm the impression that both cDNA clones code for the identical polypeptide, which is highly homologous with human lamins A and C as well as with mammalian intermediate filament proteins. The two clones differ in their 3'-untranslated regions. In situ hybridization of lamin cDNA clones to Drosophila polytene chromosomes shows only a single locus of hybridization at or near position 25F on the left arm of chromosome 2. Southern blot analyses of genomic DNA are consistent with the notion that a single or only a few highly similar genes encoding Drosophila nuclear lamin Dm0 exist in the genome.
通过使用果蝇核纤层蛋白特异性单克隆抗体筛选λ-gt11 cDNA表达文库,已鉴定出一个编码果蝇核纤层蛋白Dm1和Dm2一部分的cDNA克隆。使用初始cDNA作为探针,通过Northern印迹分析鉴定出两种不同的受发育调控的mRNA种类,并且已经分离出显然对应于每种信息的全长cDNA克隆。在pT7转录载体中对两个全长cDNA克隆进行体外转录,然后在小麦胚芽裂解物中进行体外翻译,这表明两个克隆都编码核纤层蛋白Dm0,即核纤层蛋白Dm1和Dm2的多肽前体。核苷酸序列分析证实了这两个cDNA克隆编码相同多肽的印象,该多肽与人类核纤层蛋白A和C以及哺乳动物中间丝蛋白高度同源。这两个克隆在其3'非翻译区有所不同。将核纤层蛋白cDNA克隆与果蝇多线染色体进行原位杂交,结果显示在2号染色体左臂上25F位置或其附近只有一个杂交位点。基因组DNA的Southern印迹分析与基因组中存在一个或仅几个编码果蝇核纤层蛋白Dm0的高度相似基因的观点一致。