The phenylalanine-28 is crucial for black carp RIG-I mediated antiviral signaling

Retinoic acid-inducible gene I (RIG-I) functions as a cytosolic sensor to recognize RNA products of the invading microorganisms and induce the production of type I interferons(IFNs). In this study, two RIG-I variants, named as bcRIG-Ia and bcRIG-Ib, were characterized in black carp (Mylopharyngodon piceus) respectively. RNA pulldown assay revealed that both bcRIG-Ia and bcRIG-Ib could bind to synthetic poly(I:C) and the RD domain was crucial for RNA binding of these two molecules. However, over-expression of bcRIG-Ib, but not bcRIG-Ia, induced the transcription of IFN promoter, and led to the improved antiviral activity against both spring viremia of carp virus (SVCV) and grass carp reovirus (GCRV). And knockdown of bcRIG-I dampened the transcription of bcViperin and bcIFNb in host cells. Truncation mutation and site mutation analysis identified that phenylalanine (F)- 28 was crucial for bcRIG-Ib oligomerization and its mediated IFN signaling. Interestingly, F28 was conserved among teleost RIG-Is and site mutation analysis revealed that F28 was essential for RIG-I mediated IFN signaling in the cyprinid fish. Thus, our study concludes that F28 is crucial for black carp RIG-I mediated antiviral signaling and suggests F28 is also essential for the activation of IFN signaling by RIG-Is from other teleost fish.