The structure and function of a novel G protein family regulating eukaryotic mRNA dynamics

在真核生物的蛋白质合成中，所有三个终止密码子都被一个多肽链释放因子eRF 1直接识别，从而从核糖体释放多肽链，另一个释放因子eRF 3是eRF 1与核糖体A位点结合所必需的。我们以前报道过一个GTP结合蛋白，其羧基端序列与真核细胞延伸因子EF 1 α同源，它与eRF 1形成复合物，在翻译终止中起eRF 3的作用，并且eRF 3由EF 1 α样的羧基端（C）和氨基端（N）结构域组成。本研究对eRF 3及其相关G蛋白的结构和功能进行了研究。主要研究结果如下：1）eRF 3的C结构域与eRF 1结合，而N结构域能够与多聚腺苷酸结合蛋白（PABP）结合，PABP与mRNA的poly（A）尾结合，推测是为了稳定mRNA。2)当eRF 3-N结构域与PABP之间的相互作用被消除时，由于poly（A）尾缩短的抑制，所有mRNA的衰减速率都降低。3）我们鉴定了PABP激活的多聚A核酸酶（PAN）复合物，发现eRF 3和PAN复合物竞争性结合PABP。这些结果表明，翻译终止偶联的mRNA衰变是通过eRF 3依赖性poly（A）缩短介导的。因此，eRF 3不仅在翻译终止中作为eRF 1载体蛋白，而且还作为mRNA降解机制的启动子。4）我们还研究了与eRF 3结构相关的其他G蛋白（Ski 7和eRFS）的功能，发现这些G蛋白的N结构域通常与参与mRNA降解机制的因子相互作用，以调节mRNA的稳定性。

In eukaryotic protein synthesis, all three termination codons are directly recognized by a polypeptide-chain releasing factor, eRF1,to release polypeptide chain from ribosome, and another releasing factor, eRF3,is required for eRF1 binding to ribosomal A site. We previously reported that a GTP-binding protein, whose carboxy-terminal sequence is homologous to the eukaryotic elongation factor EF1α, forms a complex with eRF1 to function as eRF3 in the translation termination and that eRF3 consists of the EF1α-like carboxy-terminal (C) and amino-terminal (N) domains. In the present studies, we investigated the structures and functions of eRF3 and its related G proteins. The major findings obtained in this study are summarized as follows.1)The C domain of eRF3 associated with eRF1,whereas the N domain was capable of binding to poly-adenylate binding protein (PABP) associating with the poly(A) tail of mRNAs presumably for their stabilization. 2)When the interaction between eRF3-N domain and PABP was abolished, decay rate of all mRNAs was decreased due to the inhibition of poly(A) tail shortening. 3)We identified a poly(A) nuclease (PAN) complex that is activated by PABP and found that eRF3 and PAN complex competitively bound to PABP. These results indicate that the translation termination-coupled mRNA decay is mediated through eRF3-dependent poly(A) shortening. Thus, eRF3 functions not only as an eRF1-carrier protein in the translation termination but also as an initiator of the mRNA degradation machinery. 4)We also investigated the functions of other G proteins (Ski7 and eRFS) structurally related to eRF3 and found that the N domains of these G proteins commonly interact with factors involved in mRNA-degradation machineries to regulate mRNA stabilization.