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Extensive alternative splicing and dual promoter usage generate Tcf-1 protein isoforms with differential transcription control properties.

机译:广泛的替代剪接和双重启动子的使用产生具有差异转录控制特性的Tcf-1蛋白同工型。

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摘要

Previously, we reported the isolation of cDNA clones representing four alternative splice forms of TCF-1, a T-cell-specific transcription factor. In the present study, Western blotting (immunoblotting) yielded a multitude of TCF-1 proteins ranging from 25-55 kDa, a pattern not simply explained from the known splice alternatives. Subsequent cDNA cloning, PCR amplification, and analysis by rapid amplification of 5' cDNA ends revealed (i) the presence of an alternative upstream promoter, which extended the known N terminus by 116 amino acids, (ii) the presence of four alternative exons, and (iii) the existence of a second reading frame in the last exon encoding an extended C terminus. Inclusion of the extended N terminus into the originally reported protein resulted in a striking similarity to the lymphoid factor Lef-1. Several of the TCF-1 isoforms, although less potent, mimicked Lef-1 in transactivating transcription through the T-cell receptor alpha-chain (TCR-alpha) enhancer. These data provide a molecular basis for the complexity of the expressed TCF-1 proteins and establish the existence of functional differences between these isoforms. Furthermore, the functional redundancy between Tcf-1 and Lef-1 explains the apparently normal TCR-alpha expression in single Tcf-1 or Lef-1 knockout mice despite the firm in vitro evidence for the importance of the Tcf/Lef site in the TCR-alpha enhancer.
机译:以前,我们报道了代表TCF-1(一种T细胞特异性转录因子)TCF-1四种替代剪接形式的cDNA克隆的分离。在本研究中,蛋白质印迹(免疫印迹)产生了25-55 kDa范围内的多种TCF-1蛋白,这种模式不能从已知的剪接选择中简单解释。随后的cDNA克隆,PCR扩增和通过5'cDNA末端的快速扩增进行的分析显示(i)存在另一个上游启动子,该启动子将已知的N末端扩展了116个氨基酸,(ii)存在四个替代外显子, (iii)在最后一个外显子中存在第二阅读框,其编码扩展的C端。最初报道的蛋白质中包含扩展的N末端导致与淋巴因子Lef-1惊人相似。几种TCF-1亚型,尽管效力较弱,但在通过T细胞受体α-链(TCR-α)增强子反式转录转录时模仿了Lef-1。这些数据为表达的TCF-1蛋白的复杂性提供了分子基础,并确定了这些同工型之间存在功能差异。此外,尽管有确凿的证据证明TCF / Lef位点在TCR中的重要性,但Tcf-1和Lef-1之间的功能冗余解释了单只Tcf-1或Lef-1基因敲除小鼠中看似正常的TCR-alpha表达。 -alpha增强剂。

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