Ii S, Sobell J L, Sommer S S
Department of Biochemistry and Molecular Biology, Mayo Clinic/Foundation, Rochester, MN 55905.
Am J Hum Genet. 1992 Jan;50(1):29-41.
Traditionally, clinical research has sought to determine the molecular basis of clinical signs and symptoms. Increasingly, the traditional process will be reversed, as many structural protein variants are elucidated as a result of powerful PCR-based methods. Herein we describe a variant of transthyretin (TTR) found by direct genomic sequencing and illustrate the utility of PASA (PCR amplification of specific alleles) in the initial characterization of such variants. TTR is an intriguing protein of unknown function, but deposition of mutant TTR produces familial amyloidotic polyneuropathy (FAP). We identify a carrier of a variant TTR in which threonine119 is changed to methionine (T119----M). T119 is invariant in five mammalian species, suggesting that this residue is important for normal protein function. To determine the frequency of the M119 variant, individuals of northern- and western-European descent were rapidly screened by generating a PASA assay for the sequence change. Four additional individuals were found to be heterozygous for the mutation, for a total of five M119 alleles in 1,666 genes (1/333). Clinical records, initial clinical interviews, and family history of these patients hint at a high frequency of early-onset venous insufficiency and perhaps mild renal dysfunction. Haplotype analysis on the heterozygotes could be performed, despite the absence of samples from relatives, by performing "double PASA." The haplotype data suggest that the M119 variant derives from a common ancestor. The putative functional deficiency caused by TTR M119 should be most marked in the homozygotes, who can be calculated to occur in 1/100,000 conceptions. If viable, these individuals may provide important clues about the physiological role of TTR. Although the nature (if any) of disease caused by TTR M119 remains to be defined, the genetic and clinical data indicate that this mutation does not cause FAP. Future family studies can determine whether the heterozygous state for TTR M119 cosegregates with a disease or trait.
传统上，临床研究一直致力于确定临床体征和症状的分子基础。随着基于强大的聚合酶链反应（PCR）方法阐明了许多结构蛋白变体，传统流程越来越多地被逆转。在此，我们描述了通过直接基因组测序发现的转甲状腺素蛋白（TTR）变体，并说明了等位基因特异性PCR扩增（PASA）在此类变体初始特征分析中的应用。TTR是一种功能未知的有趣蛋白质，但突变型TTR的沉积会导致家族性淀粉样多神经病（FAP）。我们鉴定出一名TTR变体携带者，其中苏氨酸119被替换为甲硫氨酸（T119→M）。T119在五种哺乳动物物种中是不变的，这表明该残基对正常蛋白质功能很重要。为了确定M119变体的频率，通过针对该序列变化生成PASA检测法，对北欧和西欧血统的个体进行了快速筛查。另外发现4名个体为该突变的杂合子，在1666个基因中共检测到5个M119等位基因（1/333）。这些患者的临床记录、初步临床访谈和家族史提示早发性静脉功能不全的发生率较高，可能还有轻度肾功能不全。尽管没有亲属的样本，但通过进行“双重PASA”，仍可对杂合子进行单倍型分析。单倍型数据表明M119变体源自一个共同祖先。由TTR M119引起的假定功能缺陷在纯合子中应该最为明显，据计算，纯合子的发生率为1/100,000次受孕。如果这些个体能够存活，可能会为TTR的生理作用提供重要线索。虽然由TTR M119引起的疾病性质（如果有的话）仍有待确定，但遗传和临床数据表明该突变不会导致FAP。未来的家族研究可以确定TTR M119的杂合状态是否与一种疾病或性状共分离。
