TRBP 确保了在 RNA 拥挤环境中前体 microRNA 的高效 Dicer 加工。
TRBP ensures efficient Dicer processing of precursor microRNA in RNA-crowded environments.
机构信息
Kavli Institute of NanoScience, Delft University of Technology, 2629HZ Delft, The Netherlands.
Department of BioNanoScience, Delft University of Technology, 2629HZ Delft, The Netherlands.
出版信息
Nat Commun. 2016 Dec 9;7:13694. doi: 10.1038/ncomms13694.
The RNA-binding protein TRBP is a central component of the Dicer complex. Despite a decade of biochemical and structural studies, the essential functionality of TRBP in microRNA (miRNA) biogenesis remains unknown. Here we show that TRBP is an integral cofactor for time-efficient Dicer processing in RNA-crowded environments. We competed for Dicer processing of pre-miRNA with a large amount of cellular RNA species and found that Dicer-TRBP, but not Dicer alone, remains resilient. To apprehend the mechanism of this substrate selectivity, we use single-molecule fluorescence. The real-time observation reveals that TRBP acts as a gatekeeper, precluding Dicer from engaging with pre-miRNA-like substrates. TRBP acquires the selectivity using the PAZ domain of Dicer, whereas Dicer moderates the RNA-binding affinity of TRBP for fast turnover. This coordinated action between TRBP and Dicer accomplishes an efficient way of discarding pre-miRNA-like substrates.
RNA 结合蛋白 TRBP 是 Dicer 复合物的核心组成部分。尽管经过了十年的生化和结构研究，但 TRBP 在 microRNA（miRNA）生物发生中的基本功能仍然未知。在这里，我们表明 TRBP 是在 RNA 拥挤环境中高效进行 Dicer 加工的必不可少的辅助因子。我们用大量的细胞 RNA 竞争 Dicer 对 pre-miRNA 的加工，发现 Dicer-TRBP（而非单独的 Dicer）保持稳定。为了理解这种底物选择性的机制，我们使用单分子荧光。实时观察表明，TRBP 充当了一种门控因子，阻止 Dicer 与 pre-miRNA 样底物结合。TRBP 使用 Dicer 的 PAZ 结构域获得选择性，而 Dicer 则调节 TRBP 与 RNA 的结合亲和力，以实现快速周转。TRBP 和 Dicer 之间的这种协调作用实现了一种有效的丢弃 pre-miRNA 样底物的方法。
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