Colegrove-Otero Lucy J, Devaux Agathe, Standart Nancy
Department of Biochemistry, University of Cambridge, UK.
Mol Cell Biol. 2005 Oct;25(20):9028-39. doi: 10.1128/MCB.25.20.9028-9039.2005.
Xenopus laevis Vg1 mRNA undergoes both localization and translational control during oogenesis. We previously characterized a 250-nucleotide AU-rich element, the Vg1 translation element (VTE), in the 3'-untranslated region (UTR) of this mRNA that is responsible for the translational repression. UV-cross-linking and immunoprecipitation experiments, described here, revealed that the known AU-rich element binding proteins, ElrA and ElrB, and TIA-1 and TIAR interact with the VTE. The levels of these proteins during oogenesis are most consistent with a possible role for ElrB in the translational control of Vg1 mRNA, and ElrB, in contrast to TIA-1 and TIAR, is present in large RNP complexes. Immunodepletion of TIA-1 and TIAR from Xenopus translation extract confirmed that these proteins are not involved in the translational repression. Mutagenesis of a potential ElrB binding site destroyed the ability of the VTE to bind ElrB and also abolished translational repression. Moreover, multiple copies of the consensus motif both bind ElrB and support translational control. Therefore, there is a direct correlation between ElrB binding and translational repression by the Vg1 3'-UTR. In agreement with the reporter data, injection of a monoclonal antibody against ElrB into Xenopus oocytes resulted in the production of Vg1 protein, arguing for a role for the ELAV proteins in the translational repression of Vg1 mRNA during early oogenesis.
非洲爪蟾的Vg1信使核糖核酸在卵子发生过程中经历定位和翻译控制。我们之前鉴定出该信使核糖核酸3'非翻译区(UTR)中有一个250个核苷酸的富含AU的元件,即Vg1翻译元件(VTE),它负责翻译抑制。本文所述的紫外线交联和免疫沉淀实验表明,已知的富含AU的元件结合蛋白ElrA和ElrB,以及TIA-1和TIAR与VTE相互作用。卵子发生过程中这些蛋白的水平与ElrB在Vg1信使核糖核酸翻译控制中可能发挥的作用最为一致,并且与TIA-1和TIAR不同,ElrB存在于大型核糖核蛋白复合体中。从非洲爪蟾翻译提取物中免疫去除TIA-1和TIAR证实这些蛋白不参与翻译抑制。一个潜在的ElrB结合位点的诱变破坏了VTE结合ElrB的能力,也消除了翻译抑制。此外,共有基序的多个拷贝既能结合ElrB又能支持翻译控制。因此,ElrB结合与Vg1 3'-UTR的翻译抑制之间存在直接关联。与报告基因数据一致,将抗ElrB单克隆抗体注射到非洲爪蟾卵母细胞中导致Vg1蛋白产生,这表明ELAV蛋白在卵子发生早期对Vg1信使核糖核酸的翻译抑制中发挥作用。