Saccharomyces cerevisiae Sof1p associates with 35S Pre-rRNA independent from U3 snoRNA and Rrp5p
Saccharomyces cerevisiae Sof1p associates with 35S Pre-rRNA independent from U3 snoRNA and Rrp5p
复制标题
DOI：
10.1128/ec.5.3.427-434.2006
复制
发表时间：
2006-03-01
期刊：
影响因子：
--
通讯作者：
Vos, JC
中科院分区：
文献类型：
--
作者：
Bax, R;Vos, HR;Vos, JC
Sof 1 p是一种反式作用蛋白，对酿酒酵母中40 S核糖体亚基的生物合成至关重要。由于其参与前体rRNA的早期切割步骤，与Nop 1 p的相互作用及其共沉淀U3 snoRNA的能力，Sof 1 p迄今为止被认为是U3 snoRNP特异性的蛋白质。为了确定U3 snoRNA中是否存在与Sof 1 p直接或间接相关的位点，我们研究了ProtA标记的Sof 1 p与U3 snoRNA突变体共免疫沉淀的能力。测试的突变对从细胞提取物中回收突变体U3没有显著影响。进一步的免疫共沉淀实验，使用的细胞，可以是基因耗尽的Sof 1 p或U3 snoRNA证明，这两个因素相互独立地与35 S前体RNA。事实上，在35 S前rRNA转录被阻断的细胞中，Sof 1 p和U3 snoRNA之间的关联被消除。最后，我们发现，通过基因耗竭常见的Nop 58 p蛋白质，盒C/D snoRNP水平的总体降低并不影响Sof 1 p与35 S前rRNA的共沉淀。从这些数据中，我们得出结论，Sof 1 p不组装成90 S前核糖体的一部分，U3，或任何其他框C/D，snoRNP。早期和独立组装的反式作用因子Rrp 5 p也被证明是Sof 1 p组装的关键。
Sof1p is a trans-acting protein that is essential for biogenesis of the 40S ribosomal subunits in the yeast Saccharomyces cerevisiae. Because of its involvement in the early cleavage steps of precursor rRNA, its interaction with Nop1p and its ability to coprecipitate U3 snoRNA, Sof1p has so far been regarded as a protein that is specific to the U3 snoRNP. To determine whether a site exists within U3 snoRNA with which Sof1p directly or indirectly associates, we studied the ability of ProtA-tagged Sof1p to coimmunoprecipitate mutant versions of U3 snoRNA. None of the tested mutations had a significant effect on the recovery of mutant U3 from cell extracts. Further coimmunoprecipitation experiments, using cells that could be genetically depleted for either Sof1p or U3 snoRNA demonstrated that the two factors associate independently of each other with the 35S precursor RNA. Indeed, association between Sof1p and U3 snoRNA was abolished in cells in which 35S pre-rRNA transcription was blocked. Finally, we found that an overall reduction in the levels of box C/D snoRNPs by genetic depletion of the common Nop58p protein did not affect coprecipitation of 35S pre-rRNA by Sof1p. From these data, we conclude that Sof1p does not assemble into the 90S preribosome as part of the U3, or any other box C/D, snoRNP. The early and independently assembling trans-acting factor Rrp5p also proved to be dispensable for assembly of Sof1p.