Abstract
TERRA, TElomeric Repeat-containing RNA, is a long non-coding RNA transcribed from telomeres. Emerging evidence indicates that TERRA regulates telomere maintenance and chromosome end protection in normal and cancerous cells. However, the mechanism of how TERRA contributes to telomere functions is still unclear, partially owing to the shortage of approaches to track and manipulate endogenous TERRA molecules in live cells. Here, we developed a method to visualize TERRA in live cells via a combination of CRISPR Cas13 RNA labeling and SunTag technology. Single-particle tracking reveals that TERRA foci undergo anomalous diffusion in a manner that depends on the timescale and telomeric localization. Furthermore, we used a chemically-induced protein dimerization system to manipulate TERRA subcellular localization in live cells. Overall, our approaches to monitor and control TERRA locations in live cells provide powerful tools to better understand its roles in telomere maintenance and genomic integrity.
Highlights
Telomeres, the repetitive DNA sequences at chromosome ends, are coated by the Shelterin protein complex to protect them from incorrect fusion and recombination as DNA double-strand breaks (Palm and de Lange, 2008; O’Sullivan and Karlseder, 2010; Maciejowski and De Lange, 2017)
There is a growing consensus that characteristic distributions and dynamics of TElomeric Repeat-containing RNAs (TERRAs) correlate with its function
Robust tools to track and manipulate the spatiotemporal dynamics of TERRA are vital to understanding TERRA functions
Summary
The repetitive DNA sequences at chromosome ends, are coated by the Shelterin protein complex to protect them from incorrect fusion and recombination as DNA double-strand breaks (Palm and de Lange, 2008; O’Sullivan and Karlseder, 2010; Maciejowski and De Lange, 2017). TERRAs are transcribed from the subtelomeric regions towards chromosome ends by RNA polymerase II and are highly heterogeneous transcripts with sizes ranging from 100 nt to 9 kb in mammalian cells (Azzalin et al, 2007; Schoeftner and Blasco, 2008; Bettin et al, 2019). A growing body of studies indicates that TERRA actively regulates telomere function and maintenance (Azzalin et al, 2007; Deng et al, 2009; Wang et al, 2015; Chu et al, 2017; Feretzaki et al, 2020). TERRA is involved in telomere maintenance of cancer cells (De Silanes et al, 2014; Chu et al, 2017; Bettin et al, 2019).
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