RNA abundance is tightly regulated in eukaryotic cells by modulating the kinetic rates of RNA production, processing, and degradation. To date, little is known about time-dependent kinetic rates during dynamic processes. Researchers at the Max Delbrück Center for Molecular Medicine have developed SLAM-Drop-seq, a method that combines RNA metabolic labeling and alkylation of modified nucleotides in methanol-fixed cells with droplet-based sequencing to detect newly synthesized and preexisting mRNAs in single cells. As a first application, the researchers sequenced 7280 HEK293 cells and calculated gene-specific kinetic rates during the cell cycle using the novel package Eskrate. Of the 377 robust-cycling genes that they identified, only a minor fraction is regulated solely by either dynamic transcription or degradation (6 and 4%, respectively). By contrast, the vast majority (89%) exhibit dynamically regulated transcription and degradation rates during the cell cycle. This study thus shows that temporally regulated mRNA degradation is fundamental for the correct expression of a majority of cycling genes. SLAM-Drop-seq, combined with Eskrate, is a powerful approach to understanding the underlying mRNA kinetics of single-cell gene expression dynamics in continuous biological processes.
A novel approach (SLAM-Drop-seq) is coupled to a robust computational analysis method (Eskrate) to discriminate new and preexisting transcripts in single cells and to calculate their time-dependent kinetic rates of synthesis and decay during the cell cycle.