Investigation of mechanism for the genesis of polymorphisms of the coagulation factor XIII and PGM1 genes

我们证明了凝血因子XIII“a”亚基的遗传多态性和5个外显子的新序列多态性的分子基础。发现4个F13 A等位基因是由外显子12和外显子14的3个位点的核苷酸替换定义的。我们开发了PCR-RFLPs，通过它可以在基因组水平上检测这些取代。除了这三个位点，我们的特点是新的多态性位点的外显子2，5，8，12和14在高加索人群，而这些网站是非多态性的，除了外显子2的网站在日本。这些多态性位点被发现作为单倍型传递。理论上72种可能的组合中，共有18种单倍型出现在高加索人群中，这意味着频繁的重组事件而不是重复的点突变导致单倍型的产生。此外，我们揭示了14个不同的变异等位基因和一个缺陷等位基因的序列差异，通过表征其完整的序列。14个变异等位基因的核苷酸变化导致的氨基酸取代与其产物在IEF凝胶中的迁移率变化一致。该缺失等位基因是由于F13 A基因第5内含子剪接受体位点的单核苷酸突变引起的异常剪接所致，提示F13 A基因区域存在重组热点。我们现在正在研究内含子和5 '-和3'-侧翼区的序列。

We demonstrated molecular basis of the genetic polymorphism of the coagulation factor XIII "a" subunit and novel sequence polymorphisms in five exons. Four F13A alleles were found to be defined by nucleotide substitutions at the three sites of exon 12 and exon 14. We developed PCR-RFLPs by which these substitutions can be detected at the genomic level. In addition to the three sites, we characterized novel polymorphic sites of exons 2,5,8,12 and 14 in Caucasian populations whereas these sites were non-polymorphic in Japanese except for the site of exon 2. These polymorphic sites were found to be transmitted as haplotypes. A total of 18 haplotypes out of theoretically 72 possible combinations were found to occur in the Caucasian populations, implying that frequent recombination events rather than reccurent point mutations lead to production of the haplotypes. Furthermore, we revealed sequence differences for 14 distinct variant alleles and one deficient allele by characterizing their complete sequences. Amino acid substitutions resulting from the nucleotide changes of the 14 variant alleles were consistent with mobility shift of their products in IEF gels. The deficient allele was found to result from abnormal splicing caused by a single nucleotide mutation at the splice-acceptor site in intron 5 of the F13A gene.The data obtained in this study suggest that there are recombination hot-spots in the F13A gene region. We are now investigating the sequences of introns and 5'-and 3'-flanking regions.