Identification of a Novel C-Terminal Truncated WT1 Isoform with Antagonistic Effects against Major WT1 Isoforms.
Identification of a Novel C-Terminal Truncated WT1 Isoform with Antagonistic Effects against Major WT1 Isoforms.
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DOI：
10.1371/journal.pone.0130578
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Oji Y
中科院分区：
文献类型：
--
作者：
Tatsumi N;Hojo N;Sakamoto H;Inaba R;Moriguchi N;Matsuno K;Fukuda M;Matsumura A;Hayashi S;Morimoto S;Nakata J;Fujiki F;Nishida S;Nakajima H;Tsuboi A;Oka Y;Hosen N;Sugiyama H;Oji Y
WT 1基因由10个外显子组成，编码一个锌指转录因子。有四种主要的WT 1亚型，由两个位点的选择性剪接产生，外显子5（17 AA）和外显子9（KTS）。所有主要的WT 1亚型在白血病和实体瘤中过表达，并发挥致癌作用，如抑制细胞凋亡，促进细胞增殖，迁移和侵袭。在本研究中，鉴定了一种新的选择性剪接WT 1亚型，其具有延伸的外显子4（命名为外显子4a），在3'端具有额外的153 bp（命名为4a序列），并命名为Ex 4a（+）WT 1亚型。外显子4a的插入导致在外显子4a的阅读框中引入提前翻译终止密码子，并产生C-末端截短的缺乏锌指DNA结合结构域的WT 1蛋白。过表达截短型Ex 4a（+）WT 1抑制了WT 1介导的抗凋亡Bcl-xL基因启动子的转录激活，并诱导线粒体损伤和凋亡。相反，通过Ex 4a特异性siRNA抑制Ex 4a（+）WT 1亚型会减弱细胞凋亡。这些结果表明Ex 4a（+）WT 1亚型对主要WT 1亚型的抗凋亡功能具有显性负效应。Ex 4a（+）WT 1亚型在髓性白血病和实体瘤细胞中内源性表达为次要亚型，并且在细胞凋亡期间无论主要WT 1亚型的减少如何都会增加，表明主要WT 1亚型的抗细胞凋亡功能具有显性负效应。这些结果表明Ex 4a（+）WT 1异构体具有重要的生理功能，调节主要WT 1异构体的致癌功能。
The Wilms’ tumor gene WT1 consists of 10 exons and encodes a zinc finger transcription factor. There are four major WT1 isoforms resulting from alternative splicing at two sites, exon 5 (17AA) and exon 9 (KTS). All major WT1 isoforms are overexpressed in leukemia and solid tumors and play oncogenic roles such as inhibition of apoptosis, and promotion of cell proliferation, migration and invasion. In the present study, a novel alternatively spliced WT1 isoform that had an extended exon 4 (designated as exon 4a) with an additional 153 bp (designated as 4a sequence) at the 3’ end was identified and designated as an Ex4a(+)WT1 isoform. The insertion of exon 4a resulted in the introduction of premature translational stop codons in the reading frame in exon 4a and production of C-terminal truncated WT1 proteins lacking zinc finger DNA-binding domain. Overexpression of the truncated Ex4a(+)WT1 isoform inhibited the major WT1-mediated transcriptional activation of anti-apoptotic Bcl-xL gene promoter and induced mitochondrial damage and apoptosis. Conversely, suppression of the Ex4a(+)WT1 isoform by Ex4a-specific siRNA attenuated apoptosis. These results indicated that the Ex4a(+)WT1 isoform exerted dominant negative effects on anti-apoptotic function of major WT1 isoforms. Ex4a(+)WT1 isoform was endogenously expressed as a minor isoform in myeloid leukemia and solid tumor cells and increased regardless of decrease in major WT1 isoforms during apoptosis, suggesting the dominant negative effects on anti-apoptotic function of major WT1 isoforms. These results indicated that Ex4a(+)WT1 isoform had an important physiological function that regulated oncogenic function of major WT1 isoforms.
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