Schneider David A, Nomura Masayasu
Department of Biological Chemistry, University of California, Irvine, CA 92697, USA.
Proc Natl Acad Sci U S A. 2004 Oct 19;101(42):15112-7. doi: 10.1073/pnas.0406746101. Epub 2004 Oct 11.
Previous experiments using mammalian cells suggested that after each round of transcription, RNA polymerase I (Pol I) dissociates into subunits that leave and reenter the nucleolus as individual subunits, before formation of a new initiation complex. In this study, we show that the size and subunit composition of Pol I did not change significantly when Pol I was not engaged in rRNA transcription, brought about by either the absence of Pol I-specific rDNA template or specific inhibition of the transcription initiation step that requires Rrn3p. In fact, Pol I purified from cells completely lacking rDNA repeats was more active than when purified from wild-type cells in an in vitro transcription system designed to assay active Pol I-Rrn3p complexes. Furthermore, measurements of the exchange of A135 and A190 subunits between preexistent Pol I and newly synthesized Pol I showed that these two largest subunits of Pol I do not disassociate through many rounds of transcription in vivo. Thus, Pol I is not a dynamic protein complex but rather a stable enzyme.
先前使用哺乳动物细胞进行的实验表明，在每一轮转录之后，RNA聚合酶I（Pol I）会解离成亚基，这些亚基在形成新的起始复合物之前，以单个亚基的形式离开并重新进入核仁。在本研究中，我们发现，当Pol I不参与rRNA转录时，其大小和亚基组成没有显著变化，这是由缺乏Pol I特异性rDNA模板或对需要Rrn3p的转录起始步骤进行特异性抑制所导致的。事实上，在用于检测活性Pol I-Rrn3p复合物的体外转录系统中，从完全缺乏rDNA重复序列的细胞中纯化得到的Pol I比从野生型细胞中纯化得到的Pol I更具活性。此外，对预先存在的Pol I和新合成的Pol I之间A135和A190亚基交换的测量表明，Pol I的这两个最大亚基在体内经过多轮转录后不会解离。因此，Pol I不是一个动态的蛋白质复合物，而是一种稳定的酶。
