GCRV S5 and Host EGR1 RNAs Form Chimeric RNA S5-AGUGA-EGR1: A Novel Mechanism Regulating Viral Gene Expression and Host Antiviral Response.

Viral transcripts generate chimeric RNAs through fusion with host transcripts, modulating viral infection and host immune responses. Grass carp hemorrhagic disease, caused by grass carp reovirus (GCRV), represents a severe disorder threatening grass carp aquaculture. Previous research on GCRV-host interactions focused on viral and host protein crosstalk, with chimeric RNAs rarely reported. Here, 88 non-redundant GCRV-grass carp chimeric RNAs were identified using RNA-seq. Six chimeric RNA subtypes formed by the S5 segment RNA of GCRV and the early growth response 1 (EGR1) RNA of grass carp were detected by PCR and Sanger sequencing. Sequence analysis revealed short direct repeats at two parental RNA breakpoints, with merely one copy retained in the chimeric RNA. Among six subtypes, the upstream sequence of the S5 RNA breakpoint (243-324 nt) was connected with the downstream sequence of the egr1 breakpoint (2604-3111 nt) to form the chimeric RNA S5-AGUGA-EGR1. S5-AGUGA-EGR1 formation was independent of viral RNA-dependent RNA polymerase (RdRP) yet relied on the short direct repeat AGUGA at breakpoints. Functional assays revealed that S5 RNA formed S5-AGUGA-EGR1 by acquiring a partial sequence of the 3'-untranslated region (3'-UTR) of EGR1, leading to downregulated EGR1 expression and increased viral RNA levels. Furthermore, S5-AGUGA-EGR1 promoted viral genes expressions by encoding a GCRV-grass carp chimeric protein VP5-c (a truncated VP5 with 8 amino acid residues encoded by the 3'-UTR of egr1 at the C-terminal). These findings provide an unprecedented perspective on GCRV-grass carp interactions and identify a novel mechanism underlying viral RNA-mediated regulation of host gene expression.