Inositol hexakisphosphate Functions as a Cofactor and Modulator of ADAR1 Activity

Abstract Adenosine-to-inosine (A-to-I) RNA editing by ADAR1 is a key post-transcriptional modification, and mutations in ADAR1 lead to Aicardi–Goutières syndrome (AGS), an autoimmune disorder. Despite its biological and clinical relevance, the regulation of ADAR1 activity remains incompletely understood. Using a combination of biochemical approaches, inositol-pentakisphosphate 2-kinase (IPPK)-knockout cells, molecular dynamics simulations, and a cell-permeable inositol hexakisphosphate (IP 6 ) prodrug (Pro-IP 6 ), we demonstrate that IP 6 depletion drastically reduces global RNA editing, while supplementation with Pro-IP 6 restores and even enhances editing levels. Furthermore, we identify the C6-phosphate of IP 6 as a critical determinant of ADAR1 catalytic efficiency, functioning within a hydrogen-bonding network that indirectly coordinates a Zn²⁺-ion. Finally, we show that the AGS-associated ADAR1 mutation N907S impairs RNA editing activity, most likely by altering the hydrogen-bond interaction network linking IP 6 to the ADAR1 catalytic center. Together, these findings identify IP 6 as an essential cofactor and regulator of ADAR1 activity and highlight cofactor availability and interaction networks as strategies for therapeutically modulating RNA editing.