In vivo assembly of functional U7 snRNP requires RNA backbone flexibility within the Sm-binding site
In vivo assembly of functional U7 snRNP requires RNA backbone flexibility within the Sm-binding site
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DOI：
10.1038/nsmb1075
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发表时间：
2006-04-01
影响因子：
16.8
通讯作者：
Steitz, JA
中科院分区：
文献类型：
--
作者：
Kolev, NG;Steitz, JA
后生动物中的大多数组蛋白前体 mRNA (pre-mRNA) 通过 U7 小核核糖核蛋白 (snRNP) 指导的 3' 末端切割而成熟。使用在非洲爪蟾卵母细胞中顺式切割的嵌合小鼠组蛋白 H4 前 mRNA-U7 snRNA 构建体，通过核苷酸类似物干扰作图和诱变来检查 U7 snRNP 体内组装和活性所需的 RNA 功能基团。独特的 U7 Sm 蛋白核心的组装限制了体内加工的速率，并且需要 U7 Sm 结合位点内有四个保守的核苷酸，以及 U7 末端茎环结构的正确定位和大小。令我们惊讶的是，假尿苷取代揭示了 U7 Sm 位点内特定位置对主链灵活性的要求，提供了体内生化证据，表明不寻常的 C2'-内糖构象对于 Sm 环的组装是必要的。
Most histone precursor mRNAs ( pre-mRNAs) in metazoans are matured by 3'-end cleavage directed by the U7 small nuclear ribonucleoprotein ( snRNP). RNA functional groups necessary for in vivo assembly and activity of the U7 snRNP were examined by nucleotide-analog interference mapping and mutagenesis using a chimeric mouse histone H4 pre-mRNA-U7 snRNA construct that is cleaved in cis in Xenopus laevis oocytes. Assembly of the unique U7 Sm protein core is rate limiting for processing in vivo and requires four conserved nucleotides within the U7 Sm-binding site, as well as the correct positioning and size of the U7 terminal stem-loop structure. To our surprise, pseudouridine substitution revealed a requirement for backbone flexibility at a particular position within the U7 Sm site, providing in vivo biochemical evidence that an unusual C2'-endo sugar conformation is necessary for assembly of the Sm ring.