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.

单细胞基因表达本质上是可变的，但如何控制这种可变性以响应刺激仍不清楚。在这里，我们使用单细胞 RNA-seq 和单分子 mRNA 计数 (smFISH) 来研究免疫 toll 样受体系统中的可诱导基因表达。我们表明，肿瘤坏死因子 α 的 mRNA 计数符合标准随机转换模型，而白介素 1β 的转录涉及额外的调节步骤，导致异质性增加。尽管有不同的调节模式，但对一系列基因的单细胞数据的系统分析表明，转录数的变异性受到一系列条件下的平均响应的线性限制。smFISH 计数的数学模型和染色质状态的实验扰动表明，线性约束是通过调节转录爆发以及基因特异性关系而出现的。总体而言，我们的分析表明，诱导型单细胞 mRNA 反应的变异性受到转录爆发的限制。