Two steps in Maf1-dependent repression of transcription by RNA polymerase III
Two steps in Maf1-dependent repression of transcription by RNA polymerase III
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DOI：
10.1074/jbc.m412375200
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发表时间：
2005-02-25
影响因子：
4.8
通讯作者：
Willis, IM
中科院分区：
文献类型：
--
作者：
Desai, N;Lee, JH;Willis, IM
在酿酒酵母中，Maf1 对于介导 RNA 聚合酶 (pol) III 响应不同细胞条件的转录抑制至关重要。这些条件激活不同的信号通路，这些信号通路在 Maf1 或以上汇聚。因此，Maf1 依赖性抑制被认为涉及对 pol III 机制的一组常见的下游抑制作用。在这里，我们为这一观点提供了支持，并定义了 Maf1 依赖性转录抑制的两个步骤。我们发现，氯丙嗪 (CPZ) 诱导的 pol III 转录抑制是通过抑制转录因子 (TF) IIIB 从头组装到 DNA 上以及将 pol III 招募到预组装的 TFIIIB-DNA 复合物中来实现的。此外，Brf1 被确定为 CPZ 处理细胞提取物中的抑制目标。 Maf1-Brf1 和 Maf1-pol III 相互作用与重组 Maf1 对 TFIIIB-DNA 复合物组装和聚合酶招募的抑制有关。免疫共沉淀实验证实了酵母提取物中的这些相互作用，并证明 Maf1 在抑制条件下不会差异性地隔离 Brf1 或 pol III。结果表明 Maf1 通过非化学计量机制抑制 pol III 转录。
In Saccharomyces cerevisiae, Maf1 is essential for mediating the repression of transcription by RNA polymerase (pol) III in response to diverse cellular conditions. These conditions activate distinct signaling pathways that converge at or above Maf1. Thus, Maf1-dependent repression is thought to involve a common set of downstream inhibitory effects on the pol III machinery. Here we provide support for this view and define two steps in Maf1-dependent transcriptional repression. We show that chlorpromazine (CPZ)-induced repression of pol III transcription is achieved by inhibiting de novo assembly of transcription factor (TF) IIIB onto DNA as well as the recruitment of pol III to preassembled TFIIIB-DNA complexes. Additionally Brf1 was identified as a target of repression in extracts of CPZ-treated cells. Maf1-Brf1 and Maf1-pol III interactions were implicated in the inhibition of TFIIIB-DNA complex assembly and polymerase recruitment by recombinant Maf1. Co-immunoprecipitation experiments confirmed these interactions in yeast extracts and demonstrated that Maf1 does not differentially sequester Brf1 or pol III under repressing conditions. The results suggest that Maf1 functions by a non-stoichiometric mechanism to repress pol III transcription.