PROTEIN-BINDING SITES ARE CONSERVED IN U1 SMALL NUCLEAR-RNA FROM INSECTS AND MAMMALS
PROTEIN-BINDING SITES ARE CONSERVED IN U1 SMALL NUCLEAR-RNA FROM INSECTS AND MAMMALS
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DOI：
10.1073/pnas.80.5.1217
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发表时间：
1983-01-01
期刊：
影响因子：
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通讯作者：
PEDERSON, T
中科院分区：
文献类型：
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作者：
WIEBEN, ED;MADORE, SJ;PEDERSON, T
为了深入了解小核RNA的核糖核蛋白(RNP)结构，将HeLa细胞Poly(A)+mRNA翻译到网织红细胞裂解液中，并用人自身免疫抗体检测了35S标记的蛋白质与单个小核RNA物种的体外结合。分子量为32,000的蛋白质可与U1 RNA结合，但不能与U2、U4、U5或U6结合。所产生的U1 RNP复合体可被Sm和RNP抗体识别。U2RNA还与蛋白质形成复合体，被Sm抗体识别。Mw 32,000蛋白与U2 RNA缺乏结合并不是由于U2在体外系统中未能结合特定蛋白。用果蝇Poly(A)+mRNA进行的类似的翻译组装实验表明，先前在果蝇U1 RNP中发现的一个相对分子质量为26,000的蛋白质在体外也与U1 RNA结合。当HeLa或果蝇mRNA的翻译产物与其他物种的U1 RNA一起呈现时，分子量为32,000和26,000的蛋白质识别异源U1上的结合部位，在这两种情况下，都形成了RNP抗体识别的复合体。在人类细胞中，一个相对分子质量为32,000的蛋白质是U1 RNA所特有的，该蛋白质和一个相对分子质量为26,000的同源物的U1 RNA结合位点在进化中高度保守。这些位点可能是人和果蝇U1 RNA 5‘端相同的13个核苷酸，也可能是U1二级结构的一个高度保守的方面。
To gain insight into the ribonucleoprotein (RNP) structure of small nuclear RNA, HeLa cell poly(A)+ mRNA was translated in a reticulocyte lysate, and the in vitro binding of 35S-labeled proteins to individual small nuclear RNA species was examined by using human autoimmune antibodies. A MW 32,000 protein binds to U1 RNA but not to U2, U4, U5, or U6. The resulting U1 RNP complex is recognized both by Sm and RNP antibodies. U2 RNA also forms a complex with protein, which is recognized by Sm antibody. The lack of binding of the MW 32,000 protein to U2 RNA is not due to a failure of U2 to bind specific proteins in the in vitro system. Similar translation-assembly experiments with Drosophila poly(A)+ mRNA reveal that a MW 26,000 protein identified previously in Drosophila U1 RNP also binds to U1 RNA in vitro. When the translation products of HeLa or Drosophila mRNA are presented with U1 RNA of the other species, the MW 32,000 and 26,000 proteins recognize binding sites on the heterologous U1 and, in both cases, form complexes recognized by RNP antibody. A MW 32,000 protein is unique to U1 RNA in human cells and the U1 RNA binding sites for this and a MW 26,000 homologue were highly conserved in evolution. These sites may be the identical 13 nucleotides at the 5'' ends of human and Drosophila U1 RNA or a highly conserved aspect of U1 secondary structure.