Gene-Specific Linear Trends Constrain Transcriptional Variability of the Toll-like Receptor Signaling

Highlights

• Single-cell TNF-α and IL-1β mRNA responses are differentially controlled

• Variability of TLR-induced responses scale linearly with mean mRNA counts

• Gene-specific constraints emerge via modulation of transcriptional bursting

• Chromatin state regulates transcriptional bursting of IL-1β

Summary

Single-cell gene expression is inherently variable, but how this variability is controlled in response to stimulation remains unclear. Here, we use single-cell RNA-seq and single-molecule mRNA counting (smFISH) to study inducible gene expression in the immune toll-like receptor system. We show that mRNA counts of tumor necrosis factor α conform to a standard stochastic switch model, while transcription of interleukin-1β involves an additional regulatory step resulting in increased heterogeneity. Despite different modes of regulation, systematic analysis of single-cell data for a range of genes demonstrates that the variability in transcript count is linearly constrained by the mean response over a range of conditions. Mathematical modeling of smFISH counts and experimental perturbation of chromatin state demonstrates that linear constraints emerge through modulation of transcriptional bursting along with gene-specific relationships. Overall, our analyses demonstrate that the variability of the inducible single-cell mRNA response is constrained by transcriptional bursting.