title: Single-cell NF-kappaB dynamics reveal digital activation and analogue information processing.
abstract: Cells operate in dynamic environments using extraordinary communication capabilities that emerge from the interactions of genetic circuitry. The mammalian immune response is a striking example of the coordination of different cell types. Cell-to-cell communication is primarily mediated by signalling molecules that form spatiotemporal concentration gradients, requiring cells to respond to a wide range of signal intensities. Here we use high-throughput microfluidic cell culture and fluorescence microscopy, quantitative gene expression analysis and mathematical modelling to investigate how single mammalian cells respond to different concentrations of the signalling molecule tumour-necrosis factor (TNF)-alpha, and relay information to the gene expression programs by means of the transcription factor nuclear factor (NF)-kappaB. We measured NF-kappaB activity in thousands of live cells under TNF-alpha doses covering four orders of magnitude. We find, in contrast to population-level studies with bulk assays, that the activation is heterogeneous and is a digital process at the single-cell level with fewer cells responding at lower doses. Cells also encode a subtle set of analogue parameters to modulate the outcome; these parameters include NF-kappaB peak intensity, response time and number of oscillations. We developed a stochastic mathematical model that reproduces both the digital and analogue dynamics as well as most gene expression profiles at all measured conditions, constituting a broadly applicable model for TNF-alpha-induced NF-kappaB signalling in various types of cells. These results highlight the value of high-throughput quantitative measurements with single-cell resolution in understanding how biological systems operate.
authors.affiliation: Department of Bioengineering, Stanford University, Stanford, California 94305, USA.
authors.forename: Savaş
authors.initials: S
authors.lastname: Tay
authors.forename: Jacob J
authors.initials: JJ
authors.lastname: Hughey
authors.forename: Timothy K
authors.initials: TK
authors.lastname: Lee
authors.forename: Tomasz
authors.initials: T
authors.lastname: Lipniacki
authors.forename: Stephen R
authors.initials: SR
authors.lastname: Quake
authors.forename: Markus W
authors.initials: MW
authors.lastname: Covert
mesh_terms: D016475:3T3 Cells; D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000818:Animals; D018929:Cell Culture Techniques; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002470:Cell Survival; D004305:Dose-Response Relationship, Drug; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation / Q000187:drug effects*; D057166:High-Throughput Screening Assays / Q000379:methods*; D051379:Mice; D046210:Microfluidic Analytical Techniques; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D016328:NF-kappa B / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects* / Q000502:physiology*; D013269:Stochastic Processes; D013379:Substrate Specificity; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha