Characterization of the gene encoding the human Kidd blood group urea transporter protein -: Evidence for splice site mutations in Jknull individuals
Characterization of the gene encoding the human Kidd blood group urea transporter protein -: Evidence for splice site mutations in Jknull individuals
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DOI：
10.1074/jbc.273.21.12973
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发表时间：
1998-05-22
影响因子：
4.8
通讯作者：
Bailly, P
中科院分区：
文献类型：
--
作者：
Lucien, N;Sidoux-Walter, F;Bailly, P
KIDD(JK)血型是由一种完整的膜糖蛋白携带的，它通过红细胞膜运输尿素，也存在于肾脏的直血管内皮细胞上。人类血型Kidd/尿素转运蛋白基因的外显子-内含子结构已经确定，它由11个外显子组成，分布在30kb的碱基对上。成熟蛋白由外显子4-11编码，转录起始点由外显子4上游335个碱基对的5‘-快速扩增-聚合酶链式反应确定，5’-侧翼区由-837-336位核苷酸组成，含有TATA和倒转CAAT盒以及GATA-1/SP1红系特异性顺式作用调控元件。对S的非翻译区的分析表明，两个同样丰富的4.4kb和2.0kb的红系转录本是由于使用了不同的交替多聚腺苷信号而产生的。两个无关的JK(空)个体(B.S.和L.P.)的血液Southern杂交分析没有发现明显的Kidd/尿素转运蛋白基因异常，包括5‘和3’非翻译区，这两个个体缺乏所有的JK抗原和红细胞上的JK蛋白，但基因分型为纯合的JK(B)等位基因，进一步分析表明，每个变异体都发生了不同的剪接位点突变。第一个突变影响内含子5的3‘-受体剪接位点的不变G残基(变种B.S.)，第二个突变影响内含子7的5’-供体剪接位点的不变的G残基(变种L.P.)，这两个突变分别导致外显子6和7的跳跃，通过对网织红细胞中存在的JK转录本的序列分析，在非洲爪哇卵母细胞中的表达研究表明，与野生型Kidd/尿素转运蛋白相比，剪接后的转录本编码的截短蛋白并不介导尿素的易化转运，仅在卵母细胞的质膜上不表达。这些发现为JK(空)细胞缺乏Kidd/尿素转运蛋白和尿素转运缺陷提供了合理的解释。
The Kidd (JK) blood group is carried by an integral membrane glycoprotein which transports urea through the red cell membrane and is also present on endothelial cells of the vasa recta in the kidney. The exon-intron structure of the human blood group Kidd/urea transporter gene has been determined, It is organized into 11 exons distributed over 30 kilobase pairs. The mature protein is encoded by exons 4-11, The transcription initiation site was identified by 5'-rapid amplification of cDNA ends-polymerase chain reaction at 335 base pairs upstream of the translation start point located in exon 4, The 5'-flanking region, from nucleotide -837 to -336, contains TATA and inverted CAAT boxes as well as GATA-1/SP1 erythroid-specific cis-acting regulatory elements. Analysis of the S'-untranslated region reveals that the two equally abundant erythroid transcripts of 4.4 and 2.0 kilobase pairs arise from usage of different alternative polyadenylation signals.No obvious abnormality of the Kidd/urea transporter gene, including the 5'- and 3'-untranslated regions, has been detected by Southern blot analysis of the blood of two unrelated Jk(null) individuals (B.S. and L.P.), which lacks all Jk antigens and Jk proteins on red cells, but was genotyped as homozygous for a "silent" Jk(b) allele, Further analysis indicated that different splice site mutations occurred in each variant. The first mutation affected the invariant G residue of the 3'-acceptor splice site of intron 5 (variant B.S.), while the second mutation affected the invariant G residue of the 5'-donor splice site of intron 7 (variant L.P.), These mutations caused the skipping of exon 6 and 7, respectively, as seen by sequence analysis of the Jk transcripts present in reticulocytes, Expression studies in Xenopus oocytes demonstrated that the truncated proteins encoded by the spliced transcripts did not mediate a facilitated urea transport compared with the wild type Kidd/urea transporter protein and mere not expressed on the oocyte's plasma membrane. These findings provide a rational explanation for the lack of Kidd/urea transporter protein and defect in urea transport of Jk(null) cells.