Real-time imaging of RNA polymerase I activity in living human cells

Abstract

RNA polymerase I (Pol I) synthesizes about 60% of cellular RNA by transcribing multiple copies of the ribosomal RNA gene (rDNA). The transcriptional activity of Pol I controls the level of ribosome biogenesis and cell growth. However, there is currently a lack of methods for monitoring Pol I activity in real time. Here, we develop LiveArt (live imaging-based analysis of rDNA transcription) to visualize and quantify the spatiotemporal dynamics of endogenous ribosomal RNA (rRNA) synthesis. LiveArt reveals mitotic silencing and reactivation of rDNA transcription, as well as the transcriptional kinetics of interphase rDNA. Using LiveArt, we identify SRFBP1 as a potential regulator of rRNA synthesis. We show that rDNA transcription occurs in bursts and can be altered by modulating burst duration and amplitude. Importantly, LiveArt is highly effective in the screening application for anticancer drugs targeting Pol I transcription. These approaches pave the way for a deeper understanding of the mechanisms underlying nucleolar functions.