Enhanced Complex Stability and Optimal JAK Geometry are Pivotal for a Potent Type III Interferon Response

Type I and III Interferons (IFN) constitute the host system’s first line of defense against viral infections. Although the two families use distinct extracellular receptor complexes, an identical pair of Janus kinases (JAK) activates a similar set of signal transducer and activator of transcription (STATs) through a conserved pathway. Consequently, type I and III IFNs are presumed to activate a largely overlapping set of IFN-stimulated genes (ISGs) and elicit similar biological responses. Therapeutically, type III IFNs are attractive alternatives to type I IFNs due to their innate tissue specificity and lower systemic toxicity. However, a major limitation of type III IFNs is the significantly lower potency of their physiological activities compared to type I IFNs. To evaluate the role of receptor geometry in IFN signaling, we engineered cell lines that express wild-type and mutant IFNλ receptors (IFNλR1) with rotated intracellular registers with respect to the associated JAK1. Evaluation of downstream signaling and biological activity of type III IFNs in cells with varied JAK-JAK geometries uncovered variant receptors that result in potentiation of type III IFN signaling and all downstream activities. With the combined use of a high-affinity ligand to stabilize the extracellular receptor complex, we found that the optimization of the intracellular JAK-JAK geometry enhances the type III IFN antiviral and anti-proliferative activities by 2- and 3-logs, respectively. In addition to providing a molecular basis for the observed differences in potency between type I and III IFNs, this work provides deeper insights into broader cytokine signaling mechanisms and novel blueprints for modulating cytokine functions to develop next generation antiviral and anticancer therapeutics. ### Competing Interest Statement The authors have declared no competing interest.