ADAR RNA editing enzymes are high-affinity dsRNA-binding proteins that deaminate adenosines to inosines in pre-mRNA hairpins and also exert editing-independent effects. We generated a Drosophila AdarE374A mutant strain encoding a catalytically inactive Adar with CRISPR/Cas9. We demonstrate that Adar adenosine deamination activity is necessary for normal locomotion and prevents age-dependent neurodegeneration. The catalytically inactive protein, when expressed at a higher than physiological level, can rescue neurodegeneration in Adar mutants, suggesting also editing-independent effects. Furthermore, loss of Adar RNA editing activity leads to innate immune induction, indicating that Drosophila Adar, despite being the homolog of mammalian ADAR2, also has functions similar to mammalian ADAR1. The innate immune induction in fly Adar mutants is suppressed by silencing of Dicer-2, which has a RNA helicase domain similar to MDA5 that senses unedited dsRNAs in mammalian Adar1 mutants. Our work demonstrates that the single Adar enzyme in Drosophila unexpectedly has dual functions.

• MeSH
• Adenosine Deaminase chemistry   genetics   MeSH
• Adenosine Monophosphate metabolism   MeSH
• Point Mutation genetics   MeSH
• Nerve Degeneration pathology   MeSH
• Drosophila melanogaster genetics   immunology   MeSH
• RNA Editing genetics   MeSH
• Catalysis MeSH
• Locomotion MeSH
• RNA, Messenger genetics   metabolism   MeSH
• Brain metabolism   MeSH
• Immunity, Innate genetics   MeSH
• Protein Domains MeSH
• Drosophila Proteins chemistry   genetics   metabolism   MeSH
• Gene Expression Regulation MeSH
• Ribonuclease III metabolism   MeSH
• RNA Helicases metabolism   MeSH
• Aging pathology   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Names of Substances
• Adar protein, Drosophila MeSH Browser
• Adenosine Deaminase MeSH
• Adenosine Monophosphate MeSH
• DCR-2 protein, Drosophila MeSH Browser
• RNA, Messenger MeSH
• Drosophila Proteins MeSH
• Ribonuclease III MeSH
• RNA Helicases MeSH

BACKGROUND: In fly brains, the Drosophila Adar (adenosine deaminase acting on RNA) enzyme edits hundreds of transcripts to generate edited isoforms of encoded proteins. Nearly all editing events are absent or less efficient in larvae but increase at metamorphosis; the larger number and higher levels of editing suggest editing is most required when the brain is most complex. This idea is consistent with the fact that Adar mutations affect the adult brain most dramatically. However, it is unknown whether Drosophila Adar RNA editing events mediate some coherent physiological effect. To address this question, we performed a genetic screen for suppressors of Adar mutant defects. Adar5G1 null mutant flies are partially viable, severely locomotion defective, aberrantly accumulate axonal neurotransmitter pre-synaptic vesicles and associated proteins, and develop an age-dependent vacuolar brain neurodegeneration. RESULTS: A genetic screen revealed suppression of all Adar5G1 mutant phenotypes tested by reduced dosage of the Tor gene, which encodes a pro-growth kinase that increases translation and reduces autophagy in well-fed conditions. Suppression of Adar5G1 phenotypes by reduced Tor is due to increased autophagy; overexpression of Atg5, which increases canonical autophagy initiation, reduces aberrant accumulation of synaptic vesicle proteins and suppresses all Adar mutant phenotypes tested. Endosomal microautophagy (eMI) is another Tor-inhibited autophagy pathway involved in synaptic homeostasis in Drosophila. Increased expression of the key eMI protein Hsc70-4 also reduces aberrant accumulation of synaptic vesicle proteins and suppresses all Adar5G1 mutant phenotypes tested. CONCLUSIONS: These findings link Drosophila Adar mutant synaptic and neurotransmission defects to more general cellular defects in autophagy; presumably, edited isoforms of CNS proteins are required for optimum synaptic response capabilities in the brain during the behaviorally complex adult life stage.

• Keywords
• ADAR, Autophagy, Drosophila, Neurodegeneration, RNA editing, TOR,
• MeSH
• Adenosine Deaminase genetics   metabolism   MeSH
• Autophagy * MeSH
• Drosophila melanogaster genetics   growth & development   physiology   MeSH
• Larva genetics   growth & development   physiology   MeSH
• Mutation MeSH
• Synaptic Transmission genetics   MeSH
• Drosophila Proteins genetics   metabolism   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Adar protein, Drosophila MeSH Browser
• Adenosine Deaminase MeSH
• Drosophila Proteins MeSH