Both RIG-I and MDA5 detect alphavirus replication in concentration-dependent mode

•Both RIG-I and MDA5 detect alphavirus replication.•Alphavirus-induced transcriptional shutoff affects type I IFN induction.•Sensing of alphavirus replication by RIG-I and MDA5 depends on their concentrations.•High basal level of RIG-I and MDA5 allows IFN induction by pathogenic alphaviruses.•This dependence determines the discrepancy between the in vivo and in vitro data.

Alphaviruses are a family of positive-strand RNA viruses that circulate on all continents between mosquito vectors and vertebrate hosts. Despite a significant public health threat, their biology is not sufficiently investigated, and the mechanisms of alphavirus replication and virus-host interaction are insufficiently understood. In this study, we have applied a variety of experimental systems to further understand the mechanism by which infected cells detect replicating alphaviruses. Our new data strongly suggest that activation of the antiviral response by alphavirus-infected cells is determined by the integrity of viral genes encoding proteins with nuclear functions, and by the presence of two cellular pattern recognition receptors (PRRs), RIG-I and MDA5. No type I IFN response is induced in their absence. The presence of either of these PRRs is sufficient for detecting virus replication. However, type I IFN activation in response to pathogenic alphaviruses depends on the basal levels of RIG-I or MDA5.