An Aicardi-Goutières Syndrome-Causative Point Mutation in Adar1 Gene Invokes Multiorgan Inflammation and Late-Onset Encephalopathy in Mice.

Aicardi-Goutières syndrome (AGS) is a congenital inflammatory disorder accompanied by overactivated type I IFN signaling and encephalopathy with leukodystrophy and intracranial calcification. To date, none of the mouse models carrying an AGS-causative mutation has mimicked such brain pathology. Here, we established a mutant mouse model carrying a K948N point mutation, corresponding to an AGS-causative K999N mutation, located in a deaminase domain of the Adar1 gene that encodes an RNA editing enzyme. Adar1K948N/K948N mice displayed postnatal growth retardation. Hyperplasia of splenic white pulps with germinal centers and hepatic focal inflammation were observed from 2 mo of age. Inflammation developed in the lungs and heart with lymphocyte infiltration in an age-dependent manner. Furthermore, white matter abnormalities with astrocytosis and microgliosis were detected at 1 y of age. The increased expression of IFN-stimulated genes was detected in multiple organs, including the brain, from birth. In addition, single-nucleus RNA sequencing revealed that this elevated expression of IFN-stimulated genes was commonly observed in all neuronal subtypes, including neurons, oligodendrocytes, and astrocytes. We further showed that a K948N point mutation reduced the RNA editing activity of ADAR1 in vivo. The pathological abnormalities found in Adar1K948N/K948N mice were ameliorated by either the concurrent deletion of MDA5, a cytosolic sensor of unedited transcripts, or the sole expression of active ADAR1 p150, an isoform of ADAR1. Collectively, such data suggest that although the degree is mild, Adar1K948N/K948N mice mimic multiple AGS phenotypes, including encephalopathy, which is caused by reduced RNA editing activity of the ADAR1 p150 isoform.

RNA editing at a limited number of sites is sufficient to prevent MDA5 activation in the mouse brain.

Adenosine deaminase acting on RNA 1 (ADAR1), an enzyme responsible for adenosine-to-inosine RNA editing, is composed of two isoforms: nuclear p110 and cytoplasmic p150. Deletion of Adar1 or Adar1 p150 genes in mice results in embryonic lethality with overexpression of interferon-stimulating genes (ISGs), caused by the aberrant recognition of unedited endogenous transcripts by melanoma differentiation-associated protein 5 (MDA5). However, among numerous RNA editing sites, how many RNA sites require editing, especially by ADAR1 p150, to avoid MDA5 activation and whether ADAR1 p110 contributes to this function remains elusive. In particular, ADAR1 p110 is abundant in the mouse brain where a subtle amount of ADAR1 p150 is expressed, whereas ADAR1 mutations cause Aicardi-Goutières syndrome, in which the brain is one of the most affected organs accompanied by the elevated expression of ISGs. Therefore, understanding RNA editing-mediated prevention of MDA5 activation in the brain is especially important. Here, we established Adar1 p110-specific knockout mice, in which the upregulated expression of ISGs was not observed. This result suggests that ADAR1 p150-mediated RNA editing is enough to suppress MDA5 activation. Therefore, we further created Adar1 p110/Adar2 double knockout mice to identify ADAR1 p150-mediated editing sites. This analysis demonstrated that although the elevated expression of ISGs was not observed, only less than 2% of editing sites were preserved in the brains of Adar1 p110/Adar2 double knockout mice. Of note, we found that some sites were highly edited, which was comparable to those found in wild-type mice, indicating the presence of ADAR1 p150-specific sites. These data suggest that RNA editing at a very limited sites, which is mediated by a subtle amount of ADAR1 p150, is sufficient to prevents MDA5 activation, at least in the mouse brain.