Temporal perturbation of STAT1/2 activity reveals dynamic ligand discrimination of type I interferon signaling

Abstract

SUMMARYType-I interferon (IFN-I) subtypes signal through the same IFNα receptor (IFNAR), and initiate temporal STAT1/2 activation to orchestrate innate and adaptive immunity. It remains unknown how IFNAR discriminates between subtypes (e.g., IFNα and IFNβ), and how STAT1/2 signaling is affected by time-varying inputs. Here, we utilize our microfluidic system and live-cell imaging to quantify STAT1/2 activation dynamics in a reporter fibroblast model. Population-averaged and single-cell analyses reveal distinct STAT1/2 responses to various IFNα and IFNβ inputs. Upon continuous stimulation, cells show less sensitivity but more sustained responses to IFNα over IFNβ. A short IFNα pulse induces nearly homogeneous STAT1/2 dynamics, in contrast to heterogeneous responses in IFNβ-pulsed cells. Distinct STAT1/2 refractory states emerge upon exposure to repeated IFN-I pulses, while alternating pulse stimulation reveals that IFNβ can revoke STAT1/2 refractoriness caused by IFNα, but not vice versa. These findings highlight the differences between IFNα and IFNβ signaling and how they can elicit distinct temporal cellular behaviors during viral infection.