Ribosome stalling (also known as ribosome pausing) occurs physiologically across most of the transcriptome and is known to facilitate co-translational folding of the nascent peptide chain. Prolonged stalling, however, can also lead to ribosome collisions arising due to transcript mutations or defective nascent peptides, ultimately resulting in both the transcript and the nascent polypeptide being targeted for degradation. Global increases in such events can have severe implications for cellular proteostasis and viability.
Disome-seq involves the sequencing of mRNA fragments protected by two ribosomes packed together, a product of translational stalling where an upstream elongating ribosome runs into a downstream stalled one. Mapping of these ‘disome footprints’ provides a new benchmark for the investigation of ribosomal collisions transcriptome-wide, allowing the full extent of ribosome stalling and queuing to be elucidated. While Ribo-seq can also be used to predict the locations of stalled ribosomes, disome footprints densities have been found to reveal insights which are absent from monosome footprint data. Together, these techniques offer a comprehensive view of ribosome stalling dynamics, empowering researchers to decipher the intricacies of proteostatic changes between healthy and diseased states. In-depth expertise is essential for the analysis of Disome-seq data due to the complex nature of interpreting ribosome stalling events and distinguishing them from other translation dynamics, ensuring accurate and meaningful insights are obtained.
