Mantzouridis T D, Fragoulis E G
Department of Biochemistry and Molecular Biology, School of Biological Sciences, University of Athens, Panepistemiopolis-Kouponia, 15701 Athens, Greece.
Biochim Biophys Acta. 1998 Nov 26;1443(1-2):267-73. doi: 10.1016/s0167-4781(98)00220-6.
In insects the DOPA decarboxylase gene (Ddc) is expressed in the epidermis where dopamine derivatives promote cuticle sclerotization and the central nervous system participating in the synthesis of serotonin and dopamine. The temporal course of Ddc enzyme activity during the white prepuparium and eclosion developmental stages primarily reflects this role. Here we report that a cDNA clone for l-DOPA decarboxylase (EC 4.1.1.28) has been isolated from the white puparium stage of the insect Ceratitis capitata, the cDNA sequence has been determined and it corresponds to an mRNA of 1341 base pairs. The comparison of this cDNA sequence with that of the eclosion stage which has been previously isolated in our laboratory revealed that the cDNA from the eclosion stage codes for protein consisting of 431 amino acid (aa) residues whereas that from the white prepuparium stage for protein of 425 aa residues. In addition there is a region of 33 aa residues close to the N-terminal region of the deduced protein in which marked differences in the aa composition are observed. This result was further confirmed by Northern blot analysis in which mRNAs isolated from both developmental stages were only hybridized with their respective polymerase chain reaction synthesized stage specific probes. These results suggest that during development of the insect C. capitata two different mRNAs are produced at the white prepuparium and the eclosion stages, giving rise to two protein isoforms presumably in response to different signals.
在昆虫中,多巴脱羧酶基因(Ddc)在表皮中表达,在那里多巴胺衍生物促进表皮硬化,并且在参与血清素和多巴胺合成的中枢神经系统中也有表达。在白色预蛹期和羽化发育阶段,Ddc酶活性的时间进程主要反映了这一作用。在此我们报告,已从地中海实蝇的白色蛹期分离出L - 多巴脱羧酶(EC 4.1.1.28)的一个cDNA克隆,已确定其cDNA序列,它对应于一个1341个碱基对的mRNA。将该cDNA序列与我们实验室先前分离的羽化期的cDNA序列进行比较发现,羽化期的cDNA编码由431个氨基酸(aa)残基组成的蛋白质,而白色预蛹期的cDNA编码由425个aa残基组成的蛋白质。此外,在推导蛋白质的N端区域附近有一个33个aa残基的区域,其中观察到aa组成有明显差异。通过Northern印迹分析进一步证实了这一结果,在该分析中,从两个发育阶段分离的mRNA仅与各自的聚合酶链反应合成的阶段特异性探针杂交。这些结果表明,在地中海实蝇发育过程中,在白色预蛹期和羽化期产生了两种不同的mRNA,可能是对不同信号的响应,从而产生了两种蛋白质异构体。