Loskutoff D J, Linders M, Keijer J, Veerman H, van Heerikhuizen H, Pannekoek H
Department of Molecular Biology, Free University of Amsterdam, The Netherlands.
Biochemistry. 1987 Jun 30;26(13):3763-8. doi: 10.1021/bi00387a004.
Plasminogen activator inhibitor 1 (PAI-1) is the primary inhibitor of tissue-type plasminogen activator and thus performs an essential role in the regulation of the fibrinolytic process. It is a member of a large family of serine protease inhibitors (serpins). We determined the structure of the PAI-1 gene in order to more completely investigate the relationship of PAI-1 to other serpins and, at the same time, to begin to delineate structure-function relations in PAI-1 itself. A human genomic cosmid DNA library was screened and found to contain two independent clones, each harboring the entire PAI-1 gene. Restriction site mapping, electron microscopic inspection of heteroduplexes, and nucleotide sequence analysis all demonstrate that the PAI-1 gene is approximately 12.2 kilobase pairs in length and consists of nine exons and eight introns. All intron-exon boundaries are in accord with the "GT-AG" rule, including a cryptic acceptor splice site found in intron 7. The intron-exon pattern of the PAI-1 gene is distinct from that of most other serpins except that intron 3 of PAI-1 occupies an identical position as intron E of ovalbumin. Comparison of our data with the proposed subdomain structure of serpins suggests that seven of the eight introns may occupy a nonrandom position in the gene. These introns either delineate boundaries of individual structural subdomains or are located in random coil regions of the protein. Transcription of the PAI-1 gene in cultured vascular endothelial cells results in two distinct mRNA species. Our data suggest that these two transcripts arise by alternative polyadenylation.
纤溶酶原激活物抑制剂1（PAI-1）是组织型纤溶酶原激活物的主要抑制剂，因此在纤维蛋白溶解过程的调节中发挥着重要作用。它是丝氨酸蛋白酶抑制剂（丝氨酸蛋白酶抑制剂超家族）大家族的一员。我们确定了PAI-1基因的结构，以便更全面地研究PAI-1与其他丝氨酸蛋白酶抑制剂的关系，同时开始描绘PAI-1自身的结构-功能关系。对一个人类基因组黏粒DNA文库进行筛选，发现其中包含两个独立的克隆，每个克隆都含有完整的PAI-1基因。限制性酶切位点图谱分析、异源双链体的电子显微镜检查以及核苷酸序列分析均表明，PAI-1基因长度约为12.2千碱基对，由9个外显子和8个内含子组成。所有内含子-外显子边界均符合“GT-AG”规则，包括在内含子7中发现的一个隐蔽的剪接受体位点。PAI-1基因的内含子-外显子模式与大多数其他丝氨酸蛋白酶抑制剂不同，只是PAI-1的内含子3与卵清蛋白的内含子E占据相同位置。将我们的数据与丝氨酸蛋白酶抑制剂提出的亚结构域结构进行比较表明，8个内含子中的7个可能在基因中占据非随机位置。这些内含子要么划定各个结构亚结构域的边界，要么位于蛋白质的无规卷曲区域。在培养的血管内皮细胞中，PAI-1基因的转录产生两种不同的mRNA。我们的数据表明，这两种转录本是通过可变聚腺苷酸化产生的。
