5S RNA Structure and Interaction with Transcription Factor A. 2. Ribonuclease Probe of the 7S Particle from Xenopus laevis Immature Oocytes and RNA Exchange Properties of the 7S Particle

Janet Andersen, Nicholas Delihas*, Jay S. Hanas, Cheng-Wen Wu Lee

*Corresponding author for this work

研究成果: Article同行評審

29 引文 斯高帕斯(Scopus)

摘要

The 5S RNA complexed in the 7S particle of immature Xenopus laevis oocytes was 32P labeled at its 3” end and then subjected in situ to partial digestion using ribo-nucleases T1, T2, A, and V1 in order to study the conformation of the complexed RNA and its interaction with the transcription factor A (TFIIIA). Digested samples were gel purified to retrieve 5S RNA that was still complexed with the transcription factor protein, and the cleavages in these RNAs were analyzed on sequencing gels. The RNA associated with the 7S particle is very susceptible to ribonuclease activity despite the presence of the protein. Also, the 5S RNA in the 7S particle is in a different conformation from renatured Xenopus laevis (Xlo) 5S RNA and appears to have less secondary structure predominantly in the stem that includes helices IV and V. A species of native Xlo 5S RNA which was isolated from 7S particle preparations under nondenaturing conditions revealed a conformation that was more similar to the 5S RNA in the 7S particle than to renatured 5S RNA. Comparison of data from partial ribonuclease digestions performed on renatured 5S RNA, on the native 5S RNA, and on the complexed 5S RNA allowed us to approximate sites of protein-induced structural change in the complexed 5S RNA, which may signal protein interaction domains. These sites include the 5’ side of helices III and V. In another approach to the study of 5S RNA-TFIIIA interactions, we have observed that incubation of 32P-labeled Xlo 5S RNA with 7S particles results in the incorporation of labeled RNA into 7S particles. Heterologous eukaryotic 5S RNA can also be incorporated into 7S particles. These results suggest that a generalized eukaryotic 5S RNA structure can be recognized by TFIIIA.

原文English
頁(從 - 到)5759-5766
頁數8
期刊Biochemistry
23
發行號24
DOIs
出版狀態Published - 1 一月 1984

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