An in vitro interaction between the human U3 snRNP and 28S rRNA sequences near the α-sarcin site
An in vitro interaction between the human U3 snRNP and 28S rRNA sequences near the α-sarcin site
复制标题
人 U3 snRNP 和 α-sarcin 位点附近的 28S rRNA 序列之间的体外相互作用
DOI：
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发表时间：
1988
期刊：
影响因子：
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通讯作者：
J. Steitz
中科院分区：
文献类型：
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作者：
K. A. Parker;J. P. Bruzik;J. Steitz
将含有哺乳动物 pre-rRNA 序列的模型转录物与 HeLa 细胞提取物一起孵育，用 T1 RNase 消化，并用抗 (U3)RNP 或对照抗体进行免疫沉淀。尽管跨越转录起始位点、ETS 加工位点、18S 5' 末端和 5.8S 两个末端的转录物没有产生受保护的片段，但源自人 28S rRNA 3' 结构域的两个重叠片段被特异性免疫沉淀。这些片段的序列是使用一种新技术确定的，其中[32P]标记的片段与作为内部标记的[3H]标记的总转录物进行共同指纹。免疫沉淀的片段源自人28S的核苷酸4570-4590和4575-4590并且邻近α-八叠球蛋白位点。保护最有可能涉及 U3 RNA，因为它对微球菌核酸酶提取物的预处理敏感。 U3 和该 rRNA 区域之间的互补性在从人类到酿酒酵母的物种中在系统发育上是保守的。讨论了这一发现的可能意义。
Model transcripts containing mammalian pre-rRNA sequences were incubated with a HeLa cell extract, digested with T1 RNase, and immunoprecipitated with anti-(U3)RNP or control antibodies. Two overlapping fragments derived from the 3' domain of human 28S rRNA were specifically immunoprecipitated although transcripts which spanned the transcription initiation site, the ETS processing site, the 5' end of 18S, and both termini of 5.8S yielded no protected fragments. The sequence of these fragments was determined using a novel technique in which the [32P]-labeled fragment was co-finger-printed with [3H]-labeled total transcript serving as an internal marker. The fragments immunoprecipitated derive from nucleotides 4570-4590 and 4575-4590 of human 28S and are adjacent to the alpha-sarcin site. Protection most likely involves the U3 RNA since it is sensitive to pretreatment of the extract with micrococcal nuclease. Complementarity between U3 and this rRNA region is phylogenetically conserved in species ranging from human to S. cerevisiae. The possible significance of this finding is discussed.