Caro Florence, Bercovich Natalia, Atorrasagasti Catalina, Levin Mariano J, Vázquez Martín P
Laboratorio de Biología Molecular de la Enfermedad de Chagas--INGEBI--CONICET, Departamento de Fisiología y Biología Molecular y Celular, Facultad de Ciencias Exactas y Naturales, University of Buenos Aires, Argentina.
Exp Parasitol. 2006 Jun;113(2):112-24. doi: 10.1016/j.exppara.2005.12.015. Epub 2006 Feb 7.
The members of the PUF family of RNA-binding proteins regulate the fate of mRNAs by binding to their 3'UTR sequence elements in eukaryotes. In trypanosomes, for which gene expression is polycistronic and controlled almost exclusively by post-transcriptional processes, PUF proteins could play a crucial role. We report here the complete analysis of the PUF protein family of Trypanosoma cruzi composed of 10 members. In silico analysis predicts the existence of at least three major groups within the T. cruzi family, based on their putative binding specificity. Using yeast three hybrid assays, we tested some of these predictions for TcPUF1, TcPUF3, TcPUF5, and TcPUF8 as representatives of these groups. Data mining of the T. cruzi genome led us to describe putative binding targets for the TcPUFs of the most conserved group, TcPUF1 and TcPUF2. The targets include genes for mitochondrial proteins and protein kinases. Finally, immunolocalization experiments showed that TcPUF1 is localized in multiple discrete foci in the cytoplasm supporting its proposed function.
RNA结合蛋白PUF家族的成员通过与真核生物中mRNA的3'UTR序列元件结合来调节mRNA的命运。在锥虫中，基因表达是多顺反子的，几乎完全由转录后过程控制，PUF蛋白可能发挥关键作用。我们在此报告对克氏锥虫PUF蛋白家族的完整分析，该家族由10个成员组成。基于其假定的结合特异性，计算机分析预测克氏锥虫家族中至少存在三个主要组。使用酵母三杂交试验，我们测试了作为这些组代表的TcPUF1、TcPUF3、TcPUF5和TcPUF8的一些预测。对克氏锥虫基因组的数据挖掘使我们描述了最保守组的TcPUF（即TcPUF1和TcPUF2）的假定结合靶点。这些靶点包括线粒体蛋白和蛋白激酶的基因。最后，免疫定位实验表明TcPUF1定位于细胞质中的多个离散焦点，支持其提出的功能。
