Single-cell RNA-seq reveals that lack of cell-to-cell communication impairs p53 activation

To dissect the DNA damage response at a single-cell time series, we stimulated HCT-116 cells with the DNA damaging agent doxorubicin (dox) for 0, 4, 8 and 24 hours and performed single-cell RNA-seq using the Fluidigm C1 microfluidics platform. Additionally, to evaluate whether the DNA damage response induced by dox was influenced by cell-to-cell communication, we applied a modified C1 workflow developed by Shalek and colleagues (Shalek, et al. 2014). According to this protocol, cells were captured in the C1 chip, treated with media containing dox or vehicle, and then immediately sealed in individual chambers to prevent intercellular communication for 4 hours (on-chip stimulation, OCS). Following incubation, samples were processed for scRNA-seq as for the previous timecourse experiment. Overall design: HCT-116 cells were treated with doxorubicin (dox) for 0, 4, 8 and 24 hours and performed single-cell RNA-seq using the Fluidigm C1 microfluidics platform. To model response in absence of paracrine signaling, HCT-116 were also treated with doxorubicin via on-chip stimulation (OCS) and single-cell RNA-seq using the Fluidigm C1 microfluidics performed at the 4h timepoint.