Single-cell RNA-seq reveals dynamic paracrine control of cellular variation

Abstract

High-throughput single-cell transcriptomics offers an unbiased approach for understanding the extent, basis, and function of gene expression variation between seemingly identical cells. Here, we sequence single-cell RNA-Seq libraries prepared from over 1,700 primary mouse bone marrow derived dendritic cells (DCs) spanning several experimental conditions. We find substantial variation between identically stimulated DCs, in both the fraction of cells detectably expressing a given mRNA and the transcript’s level within expressing cells. Distinct gene modules are characterized by different temporal heterogeneity profiles. In particular, a “core” module of antiviral genes is expressed very early by a few “precocious” cells, but is later activated in all cells. By stimulating cells individually in sealed microfluidic chambers, analyzing DCs from knockout mice, and modulating secretion and extracellular signaling, we show that this response is coordinated via interferon-mediated paracrine signaling. Surprisingly, preventing cell-to-cell communication also substantially reduces variability in the expression of an early-induced “peaked” inflammatory module, suggesting that paracrine signaling additionally represses part of the inflammatory program. Our study highlights the importance of cell-to-cell communication in controlling cellular heterogeneity and reveals general strategies that multicellular populations use to establish complex dynamic responses.

Single‐cell technologies to study the immune system

“…Both the huge cell‐to‐cell variability and the amount of data these new technologies have brought to light constitute a great resource and at the same time a challenge for scientists. The parallel development of more sophisticated algorithms, faster computational approaches and new data visualization methods has already allowed scientists to gain new insights into immune system diversity 18, 23, 55, 56. Despite this incredible progress, there are still some issues that need to be solved and most of the works involving single‐cell sequencing are methodological.…”

“…An scRNA‐seq based study of bone marrow‐derived dendritic cell activation revealed that certain genes had a bimodal pattern of gene expression 55. Subsequent studies showed that paracrine signalling of a subset of fast‐responding dendritic cells affects the whole population 56. Future studies on other specific groups of immune cell populations (e.g.…”

“…2). The report from Shalek et al 55, 56. showed that the predominant isoform of several genes differs between different dendritic cell populations.…”

Computer vision for image-based transcriptomics

“…The efficiency for detecting single transcript molecules has been estimated to be 30% [13,16] and 3% [15] respectively, which is much lower than the 85% of hybridization efficiency in sm-FISH [4,17]. Such low efficiencies currently prevent these alternative methods from surveying the transcriptome with singlemolecule sensitivity and resolution in situ [18,19].…”

RNA-Seq unveils new attributes of the heterogeneous Salmonella-host cell communication

“…24 These authors sequenced single-cell RNA-Seq libraries from over 1,700 primary dendritic cells exposed to several experimental conditions, involving different stimuli and incubation times. Important messages of the study are that a few precocious cells react at early times to the stimulus; that variations exist in the fraction of cells expressing a defined mRNA and its relative levels; that expression profiles vary overtime; and, that stimulation of bystander cells occurs essentially by paracrine effects.…”