Rrn3 becomes inactivated in the process of ribosomal DNA transcription
Rrn3 becomes inactivated in the process of ribosomal DNA transcription
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DOI：
10.1074/jbc.m301093200
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发表时间：
2003-05-23
影响因子：
4.8
通讯作者：
Rothblum, LI
中科院分区：
文献类型：
--
作者：
Hirschler-Laszkiewicz, I;Cavanaugh, AH;Rothblum, LI
酵母 Rrn3 的人类同源物是一种 72 kDa 的蛋白质，对于核糖体 DNA (rDNA) 转录至关重要。尽管 Rrn3 功能在 rDNA 转录中的重要性已得到充分证实，但其作用机制尚未确定。有人认为，酵母 RNA 聚合酶 I 或哺乳动物 Rrn3 的磷酸化可调节 RNA 聚合酶 I.Rrn3 复合物的形成，该复合物可以与特定模板相互作用。这些和其他报道的差异将对 Rrn3 在转录中发挥作用的机制产生影响。例如，在酵母 rDNA 转录系统中，Rrn3 可能起催化作用，但在哺乳动物系统中，它可能起化学计量作用。因此，我们研究了 Rrn3 是否发挥催化作用或化学计量作用的问题。我们报告哺乳动物 Rrn3 成为转录反应进行时的限制因子。此外，我们证明 Rrn3 在转录反应过程中失活。例如，从已进行转录的反应中分离出的 Rrn3 不能在后续反应中激活转录。我们还表明，这种失活的 Rrn3 不仅与 RNA 聚合酶 I 解离，而且不能与 RNA 聚合酶 I 形成稳定的复合物。我们的结果表明，Rrn3 在 rDNA 转录中按化学计量发挥作用，并且其与 RNA 聚合酶 I 结合的能力在转录后丧失。
The human homologue of yeast Rrn3, a 72-kDa protein, is essential for ribosomal DNA (rDNA) transcription. Although the importance of Rrn3 function in rDNA transcription is well established, its mechanism of action has not been determined. It has been suggested that the phosphorylation of either yeast RNA polymerase I or mammalian Rrn3 regulates the formation of RNA polymerase I.Rrn3 complexes that can interact with the committed template. These and other reported differences would have implications with respect to the mechanism by which Rrn3 functions in transcription. For example, in the yeast rDNA transcription system, Rrn3 might function catalytically, but in the mammalian system it might function stoichiometrically. Thus, we examined the question as to whether Rrn3 functions catalytically or stoichiometrically. We report that mammalian Rrn3 becomes the limiting factor as transcription reactions proceed. Moreover, we demonstrate that Rrn3 is inactivated during the transcription reactions. For example, Rrn3 isolated from a reaction that had undergone transcription cannot activate transcription in a subsequent reaction. We also show that this inactivated Rrn3 not only dissociates from RNA polymerase I, but is not capable of forming a stable complex with RNA polymerase I. Our results indicate that Rrn3 functions stoichiometrically in rDNA transcription and that its ability to associate with RNA polymerase I is lost upon transcription.