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酿酒酵母PRP21基因产物是前剪接体的一个组成成分。

The Saccharomyces cerevisiae PRP21 gene product is an integral component of the prespliceosome.

作者信息

Arenas J E, Abelson J N

机构信息

Division of Biology, California Institute of Technology, Pasadena 91125.

出版信息

Proc Natl Acad Sci U S A. 1993 Jul 15;90(14):6771-5. doi: 10.1073/pnas.90.14.6771.

DOI:10.1073/pnas.90.14.6771
PMID:8341697
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC47014/
Abstract

In Saccharomyces cerevisiae, the prp21 mutation causes accumulation of unspliced pre-mRNA at the nonpermissive temperature. We have cloned the PRP21 gene by complementation of its temperature-sensitive phenotype and found it to be the same as SPP91, an extragenic suppressor of the prp9 mutation previously studied in vivo by Chapon and Legrain [Chapon, C. & Legrain, P. (1992) EMBO J. 11, 3279-3288]. We have analyzed the effects of the prp21 mutation on splicing in vitro and have found that PRP21 is a splicing factor required for prespliceosome assembly. We also have analyzed the interaction of PRP21 with splicing complexes using anti-PRP21 antibodies and found that the RNA components of the prespliceosome--U1 and U2 small nuclear RNA (snRNA) particles and pre-mRNA--are specifically coimmunoprecipitated under splicing conditions in the presence of 0.2 M KCl. At higher KCl concentrations, U1 snRNP dissociates from splicing complexes; nevertheless, U2 snRNA and pre-mRNA are still efficiently immunoprecipitated. Immunoprecipitation of both U1 and U2 snRNA as well as pre-mRNA is ATP-dependent and requires a pre-mRNA capable of supporting prespliceosome assembly. Analysis of the unbound complexes in native gels confirmed that prespliceosomes are specifically immunoprecipitated by anti-PRP21 antibodies. These results demonstrate that PRP21 is an integral component of the prespliceosome and establishes a stable interaction with U2 snRNP and/or pre-mRNA in that complex.

摘要

在酿酒酵母中，prp21突变会导致在非允许温度下未剪接的前体mRNA积累。我们通过互补其温度敏感表型克隆了PRP21基因，发现它与SPP91相同，SPP91是先前由Chapon和Legrain在体内研究的prp9突变的基因外抑制子[Chapon, C. & Legrain, P. (1992) EMBO J. 11, 3279 - 3288]。我们分析了prp21突变对体外剪接的影响，发现PRP21是前剪接体组装所需的剪接因子。我们还使用抗PRP21抗体分析了PRP21与剪接复合体的相互作用，发现在0.2 M KCl存在的剪接条件下，前剪接体的RNA成分——U1和U2小核RNA（snRNA）颗粒以及前体mRNA——会被特异性共免疫沉淀。在较高的KCl浓度下，U1 snRNP从剪接复合体中解离；然而，U2 snRNA和前体mRNA仍然能被高效免疫沉淀。U1和U2 snRNA以及前体mRNA的免疫沉淀是ATP依赖的，并且需要能够支持前剪接体组装的前体mRNA。在天然凝胶中对未结合复合体的分析证实，前剪接体被抗PRP21抗体特异性免疫沉淀。这些结果表明PRP21是前剪接体的一个组成部分，并在该复合体中与U2 snRNP和/或前体mRNA建立了稳定的相互作用

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/069efe529663/pnas01471-0406-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/14ffc5b5b82d/pnas01471-0403-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/72a385550cef/pnas01471-0404-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/3333c163f2c1/pnas01471-0405-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/cffbfa02b088/pnas01471-0405-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/069efe529663/pnas01471-0406-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/14ffc5b5b82d/pnas01471-0403-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/72a385550cef/pnas01471-0404-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/3333c163f2c1/pnas01471-0405-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/cffbfa02b088/pnas01471-0405-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4a24/47014/069efe529663/pnas01471-0406-a.jpg

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