Orntoft T F, Vestergaard E M, Holmes E, Jakobsen J S, Grunnet N, Mortensen M, Johnson P, Bross P, Gregersen N, Skorstengaard K, Jensen U B, Bolund L, Wolf H
Department of Clinical Biochemistry, Skejby University Hospital, DK 8200 Aarhus N, Denmark.
J Biol Chem. 1996 Dec 13;271(50):32260-8. doi: 10.1074/jbc.271.50.32260.
Fucosylated glycoproteins carrying alpha1-4 fucose residues are of importance for cell adhesion and as tumor markers. The Lewis gene, FUT3, encodes the only known alpha1-4-fucosyltransferase (FucT), and individuals who are deficient in this enzyme type as Lewis-negative on erythrocytes. We examined the mutational spectrum of the Lewis gene in Denmark and found 6 different mutations. Five, T59G, T202C, C314T, G508A, and T1067A, were frequent, and one, C445A, was only detected in one out of 40 individuals. Allele-specific polymerase chain reaction as well as cloning of FUT3 alleles showed that the 202 and 314 mutations were co-located on the same allele. COS7 cells transfected with an allele having the 202/314 mutations lacked enzyme activity. Polymerase chain reaction-cleavage assays were established for the genotyping of healthy individuals as well as 20 genuine Lewis-negative cancer patients and 10 non-genuine. The latter have Lewis-negative erythrocytes but saliva alpha1-4FucT activity. The genuine Lewis-negative individuals had mutations on both FUT3 alleles. In 66 healthy individuals, a gene dosage effect was detected as FUT3 heterozygous individuals had a lower alpha1-4FucT activity in saliva than did homozygous wild-type individuals. The lower enzyme level in heterozygous individuals resulted in a significantly (p < 0.04) lower level of circulating sialyl-Lewis a structure in serum. This has the clinical impact that cut-off levels in tumor marker assays should be defined on the basis of genotyping. In the group of non-genuine Lewis-negative cancer patients, whose erythrocytes convert from Lewis-positive to Lewis-negative during the disease, FUT3 heterozygosity was significantly (p < 0.05) more common.
携带α1-4岩藻糖残基的岩藻糖基化糖蛋白对于细胞黏附以及作为肿瘤标志物具有重要意义。Lewis基因，即FUT3，编码唯一已知的α1-4-岩藻糖基转移酶（FucT），缺乏这种酶类型的个体在红细胞上表现为Lewis阴性。我们研究了丹麦人群中Lewis基因的突变谱，发现了6种不同的突变。其中5种，即T59G、T202C、C314T、G508A和T1067A较为常见，而C445A仅在40名个体中的1名中检测到。等位基因特异性聚合酶链反应以及FUT3等位基因的克隆表明，202和314突变位于同一等位基因上。用具有202/314突变的等位基因转染的COS7细胞缺乏酶活性。建立了聚合酶链反应-切割试验，用于健康个体以及20名真正的Lewis阴性癌症患者和10名非真正的Lewis阴性患者的基因分型。后者的红细胞为Lewis阴性，但唾液中具有α1-4FucT活性。真正的Lewis阴性个体在两个FUT3等位基因上都有突变。在66名健康个体中，检测到基因剂量效应，因为FUT3杂合个体唾液中的α1-4FucT活性低于纯合野生型个体。杂合个体中较低的酶水平导致血清中循环唾液酸化Lewis a结构的水平显著降低（p < 0.04）。这具有临床意义，即肿瘤标志物检测的临界值应根据基因分型来确定。在非真正的Lewis阴性癌症患者组中，其红细胞在疾病过程中从Lewis阳性转变为Lewis阴性，FUT3杂合性显著更常见（p < 0.05）。
