Shared and distinct circadian and molecular effects of ADAR2 deficiency and the Gria2 R/R rescue allele

ADAR2-mediated adenosine-to-inosine RNA editing of Gria2 pre-mRNA generates an arginine codon at the Q/R site, producing the GluA2(R) subunit that confers Ca²⁺ impermeability on AMPA receptors. The survival of Adar2 ^−/− mice is ensured by the insertion of the Gria2 ^R/R allele. Therefore, phenotype associated with ADAR2 deficiency requires distinction from features shared by mice carrying this rescue allele. We compared background wild-type C57BL/6J (Bl6), Gria2 ^R/R and Adar2 ^−/− / Gria2 ^R/R mice using locomotor activity recording, light-pulse-induced phase shifting, immunohistochemistry, RNA sequencing of suprachiasmatic nucleus (SCN) at circadian time 5 (CT5) and CT15, and mass spectrometry-based proteomics of frontal-cortex samples at CT15. Free-running period was similar across genotypes, whereas Gria2 ^R/R mice displayed reduced locomotor-rhythm amplitude. Adar2 ^−/− mice showed a larger phase delay after a CT15 light pulse than Gria2 ^R/R mice, accompanied by stronger early SCN pERK1/2 immunoreactivity but comparable c-FOS induction. RNA sequencing of SCN samples identified 18, 50 and 53 CT5–CT15 differentially expressed genes in Bl6, Gria2 ^R/R and Adar2 ^−/− mice, respectively, with six genes shared across all genotypes. Most genotype-associated transcriptional differences from Bl6 were shared by the two modified lines: 149 phase-persistent differentially expressed genes were common to both contrasts, whereas direct comparison identified only one gene at CT5 and three at CT15. Proteomics analysis of frontal cortex showed a parallel pattern, with 65 shared proteins and 15 direct differences. These findings associate the broad molecular divergence from Bl6 with the genomically encoded Gria2 ^R/R allele and/or shared genetic background, while linking ADAR2 deficiency specifically to enhanced acute photic signalling and circadian resetting.