An Alternative Transcript Derived from the Trio Locus Encodes a Guanosine Nucleotide Exchange Factor with Mouse Cell-transforming Potential*
An Alternative Transcript Derived from the Trio Locus Encodes a Guanosine Nucleotide Exchange Factor with Mouse Cell-transforming Potential*
复制标题
DOI：
10.1074/jbc.m406082200
复制
发表时间：
2004-10
影响因子：
4.8
通讯作者：
N. Yoshizuka;R. Moriuchi;Tsuyoshi Mori;Kenji Yamada;S. Hasegawa;T. Maeda;T. Shimada;Yasuaki Yamada;S. Kamihira;M. Tomonaga;S. Katamine
中科院分区：
文献类型：
--
作者：
N. Yoshizuka;R. Moriuchi;Tsuyoshi Mori;Kenji Yamada;S. Hasegawa;T. Maeda;T. Shimada;Yasuaki Yamada;S. Kamihira;M. Tomonaga;S. Katamine
通过筛选来自成人T细胞白血病的新鲜白血病细胞的cDNA表达文库转化鼠成纤维细胞NIH 3 T3的潜力，我们已经确定了一种新的转化基因，命名为Tgat。Tgat在NIH 3 T3中的表达导致接触抑制的丧失、饱和密度的增加、在半固体培养基中的锚定非依赖性生长、在裸鼠中的致瘤性以及侵袭性的增加。序列比较显示，Trio基因的选择性RNA剪接参与了Tgat的产生。Tgat cDNA编码由多功能蛋白TRIO的Rho-鸟苷核苷酸交换因子（GEF）结构域和独特的C-末端15个氨基酸序列组成的蛋白产物，其分别来自Trio基因的外显子38-46和位于其最后一个外显子（外显子58）下游的新外显子。缺乏C-末端编码区的Tgat突变体cDNA保留了Rho-GEF活性，但失去了转化潜力，表明独特序列的不可或缺的作用。另一方面，治疗Tgat转化的NIH 3 T3细胞与Y-27632，Rho相关激酶的药理学抑制剂，废除其转化表型，表明Rho-GEF活性的共同参与。因此，选择性RNA剪接导致具有Rho-GEF结构域和独特的15个氨基酸的融合蛋白，是产生新癌基因Tgat的机制。
By screening cDNA expression libraries derived from fresh leukemic cells of adult T-cell leukemia for the potential to transform murine fibroblasts, NIH3T3, we have identified a novel transforming gene, designated Tgat. Expression of Tgat in NIH3T3 resulted in the loss of contact inhibition, increase of saturation density, anchorage-independent growth in a semisolid medium, tumorigenicity in nude mice, and increased invasiveness. Sequence comparison revealed that an alternative RNA splicing of the Trio gene was involved in the generation of Tgat. The Tgat cDNA encoded a protein product consisting of the Rho-guanosine nucleotide exchange factor (GEF) domain of a multifunctional protein, TRIO, and a unique C-terminal 15-amino acid sequence, which were derived from the exons 38-46 of the Trio gene and a novel exon located downstream of its last exon (exon 58), respectively. A Tgat mutant cDNA lacking the C-terminal coding region preserved Rho-GEF activity but lost the transforming potential, indicating an indispensable role of the unique sequence. On the other hand, treatment of Tgat-transformed NIH3T3 cells with Y-27632, a pharmacological inhibitor of Rho-associated kinase, abrogated their transforming phenotypes, suggesting the coinvolvement of Rho-GEF activity. Thus, alternative RNA splicing, resulting in the fusion protein with the Rho-GEF domain and the unique 15 amino acids, is the mechanism generating the novel oncogene, Tgat.