Némoz G, Zhang R, Sette C, Conti M
Department of Gynecology and Obstetrics, Stanford University School of Medicine, CA 94305, USA.
FEBS Lett. 1996 Apr 8;384(1):97-102. doi: 10.1016/0014-5793(96)00300-6.
To determine whether the expression of different PDE4D variants is unique to the rat or conserved through evolution, we have characterized the different PDE4D mRNAs expressed in human peripheral blood mononuclear cells. RT-PCR was performed using primers based on rat sequences and mRNAs from mononuclear cells. The specifically amplified fragments had a size identical to that predicted for rat PDE4D1, PDE4D2 and PDE4D3. Sequencing confirmed that these fragments are derived from the human PDE4D gene. Their sequence was highly homologous to that reported for the rat variants. cDNAs corresponding to the entire ORF of human PDE4D2 and PDE4D3 were expressed in mammalian cells, causing a large increase in PDE activity. Western blot analysis of human peripheral blood mononuclear cell extracts demonstrated the presence of proteins corresponding to the recombinant PDE4D1 and PDE4D2. The pattern of splicing and different promoter usage of the PDE4D gene is therefore conserved during evolution, which indicates an important physiological role.
为了确定不同PDE4D变体的表达是大鼠特有的还是在进化过程中保守的，我们对人类外周血单核细胞中表达的不同PDE4D mRNA进行了表征。使用基于大鼠序列的引物和来自单核细胞的mRNA进行RT-PCR。特异性扩增的片段大小与大鼠PDE4D1、PDE4D2和PDE4D3预测的大小相同。测序证实这些片段来源于人类PDE4D基因。它们的序列与报道的大鼠变体高度同源。对应于人类PDE4D2和PDE4D3整个开放阅读框的cDNA在哺乳动物细胞中表达，导致PDE活性大幅增加。对人类外周血单核细胞提取物的蛋白质印迹分析表明存在与重组PDE4D1和PDE4D2相对应的蛋白质。因此，PDE4D基因的剪接模式和不同启动子的使用在进化过程中是保守的，这表明其具有重要的生理作用。
