Elliott D J, Bourgeois C F, Klink A, Stévenin J, Cooke H J
Medical Research Council Human Genetics Unit, Western General Hospital, Crewe Road, Edinburgh EH4 2XU, Scotland.
Proc Natl Acad Sci U S A. 2000 May 23;97(11):5717-22. doi: 10.1073/pnas.97.11.5717.
RNA-binding motif (RBM) genes are found on all mammalian Y chromosomes and are implicated in spermatogenesis. Within human germ cells, RBM protein shows a similar nuclear distribution to components of the pre-mRNA splicing machinery. To address the function of RBM, we have used protein-protein interaction assays to test for possible physical interactions between these proteins. We find that RBM protein directly interacts with members of the SR family of splicing factors and, in addition, strongly interacts with itself. We have mapped the protein domains responsible for mediating these interactions and expressed the mouse RBM interaction region as a bacterial fusion protein. This fusion protein can pull-down several functionally active SR protein species from cell extracts. Depletion and add-back experiments indicate that these SR proteins are the only splicing factors bound by RBM which are required for the splicing of a panel of pre-mRNAs. Our results suggest that RBM protein is an evolutionarily conserved mammalian splicing regulator which operates as a germ cell-specific cofactor for more ubiquitously expressed pre-mRNA splicing activators.
RNA结合基序（RBM）基因存在于所有哺乳动物的Y染色体上，并与精子发生有关。在人类生殖细胞中，RBM蛋白与前体mRNA剪接机制的成分显示出相似的核分布。为了研究RBM的功能，我们使用了蛋白质-蛋白质相互作用分析来测试这些蛋白质之间可能的物理相互作用。我们发现RBM蛋白直接与剪接因子SR家族的成员相互作用，此外，还与自身强烈相互作用。我们已经绘制了负责介导这些相互作用的蛋白质结构域，并将小鼠RBM相互作用区域表达为细菌融合蛋白。这种融合蛋白可以从细胞提取物中拉下几种功能活跃的SR蛋白种类。缺失和回补实验表明，这些SR蛋白是RBM结合的唯一剪接因子，是一组前体mRNA剪接所必需的。我们的结果表明，RBM蛋白是一种进化上保守的哺乳动物剪接调节因子，作为一种生殖细胞特异性辅因子，作用于更广泛表达的前体mRNA剪接激活剂。
