Rio2p, an evolutionarily conserved, low abundant protein kinase essential for processing of 20 S Pre-rRNA in Saccharomyces cerevisiae
Rio2p, an evolutionarily conserved, low abundant protein kinase essential for processing of 20 S Pre-rRNA in Saccharomyces cerevisiae
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DOI：
10.1074/jbc.m300759200
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发表时间：
2003-06-20
影响因子：
4.8
通讯作者：
Raué, HA
中科院分区：
文献类型：
--
作者：
Geerlings, TH;Faber, AW;Raué, HA
Saccharomyces cerevisiae Rio2p（由开放阅读框Ynl207w编码）是一种功能未知的重要蛋白，与Rio1p/ Rrp10p具有显著的序列相似性。后者最近被证明是一种进化上保守的，主要是细胞质丝氨酸/苏氨酸激酶，其存在是将20s前rRNA转化为成熟的18s rRNA的D位点最终切割所必需的。通过数据库搜索，在多种真核生物和古细菌中发现了Rio2p的同源物。详细的序列比较和使用重组蛋白的体外激酶分析表明，Rio2p定义了一个与Rio2p定义的蛋白激酶相关的亚家族，但在结构和功能上都不同于Rio2p定义的蛋白激酶。在含有GAL- rio2基因的细胞中未能耗尽Rio2p，并通过Western blotting直接分析Rio2p水平，表明该蛋白含量较低。利用一个携带点突变降低激酶活性的GAL- rio2基因，我们发现由于抑制了细胞质中20s前rRNA在d位点的裂解，该突变蛋白的缺失阻断了18s rRNA的产生，而大亚基rRNA的产生不受影响。因此，Rio2p是第二个对D位点切割至关重要的蛋白激酶，也是第一个将加工缺陷与其酶活性联系起来的蛋白激酶。然而，与Rio1p/ Rrp10p相反，Rio2p似乎主要定位于细胞核中。
Saccharomyces cerevisiae Rio2p ( encoded by open reading frame Ynl207w) is an essential protein of unknown function that displays significant sequence similarity to Rio1p/ Rrp10p. The latter was recently shown to be an evolutionarily conserved, predominantly cytoplasmic serine/ threonine kinase whose presence is required for the final cleavage at site D that converts 20 S pre- rRNA into mature 18 S rRNA. A data base search identified homologs of Rio2p in a wide variety of eukaryotes and Archaea. Detailed sequence comparison and in vitro kinase assays using recombinant protein demonstrated that Rio2p defines a subfamily of protein kinases related to, but both structurally and functionally distinct from, the one defined by Rio1p. Failure to deplete Rio2p in cells containing a GAL- rio2 gene and direct analysis of Rio2p levels by Western blotting indicated the protein to be low abundant. Using a GAL- rio2 gene carrying a point mutation that reduces the kinase activity, we found that depletion of this mutant protein blocked production of 18 S rRNA due to inhibition of the cleavage of cytoplasmic 20 S pre- rRNA at site D. Production of the large subunit rRNAs was not affected. Thus, Rio2p is the second protein kinase that is essential for cleavage at site D and the first in which the processing defect can be linked to its enzymatic activity. Contrary to Rio1p/ Rrp10p, however, Rio2p appears to be localized predominantly in the nucleus.